Morphology of the Tongue Dorsal Surface of GiltheadSeabream (Sparus aurata)F. ABBATE,* M.C. GUERRERA, G. MONTALBANO, E. CIRIACO, AND A. GERMANADepartment of Morphology, Physiology, Biochemistry and Animal Productions, Faculty of Veterinary Medicine,University of Messina, Messina, Italy

KEY WORDS morphology; tongue; scanning electron microscopy; seabream; light microscopy

ABSTRACT The gilthead seabream is a food fish, one of the most frequently used in aquacul-ture. In the species of commercial interest, feeding in captivity is very important and this processis strictly related to the morphological characteristics of the oral cavity. The aim of this study is,using the standard procedures for light and scanning electron microscopy, to analyze the morphol-ogy of the tongue dorsal surface to show if relationships are present between the tongue morphol-ogy and the nutritional habits and choices of this farmed species. The main characteristic of thegilthead seabream oral cavity floor is the presence of an apical pouch, with, probably, a protectiverole mainly for the apical, free part of the tongue. Three zones, like in other teleosts, an apex, abody and a root, can be clearly distinguished. In the pouch foliate-like papillae were observed,while the whole tongue is characterized by the presence of two types of papillae, respectively witha fungiform and cylindroid aspect, both randomly distributed throughout the whole dorsal surfaceof the tongue. Scattered and numerous taste buds, with the typical pear-onion shape, together withsmall and numerous taste pores are also present, distributed throughout the tongue surface. Ourresults demonstrate that in the gilthead seabream important mechanic and sensory roles are car-ried out by specific anatomical structures. Our anatomical data could give, together with furtherbiochemical and physiological data, an important support with the aim of improving the nutritionof aquaculture species. Microsc. Res. Tech. 00:000–000, 2012. VVC 2012 Wiley Periodicals, Inc.

INTRODUCTION

The morphology of the fish oral cavity has beenwidely studied in the last decades (for review, see Horn,1998; Kapoor and Khanna, 1994; Kapoor et al., 1975).Research on feeding performance has mostly focused onmeasuring individual underlying components such assuction pressure, flow velocity, ram, or the effects of suc-tion-induced forces on prey movement during feeding(Holzman et al., 2011, 2012a,b). Considerable effortshave been directed at understanding the functionallinks between evolutionary changes in the feedingmechanism, feeding performance, and diet (Hulseyet al., 2010; Price et al., 2010). Some data are presentabout the role of food uptake, the intraoral mechanicsand the food swallowing analyzed with particularregard to the close relationships among the adaptationof fishes to different environments and the modifica-tions occurring in the oral cavity especially on thetongue anatomy (Atema, 1971; Caprio et al., 1993;Ezeasor, 1982; Fishelson and Delarea, 2004; Fishelsonet al., 2004; Hansen et al., 2002; Maina, 2000; Meyer-Rochow, 1981). Moreover, the correlations among differ-ent food resources, available in various ecological sys-tems, and the morphological modifications wererecently studied (Collar et al., 2009; Price et al., 2010;Yashpal et al., 2006, 2009). In literature some data arealso available demonstrating the numerous differencesin the fish oral cavity, particularly in the dorsal tonguesurface in the teleosts species, also regarding their feed-ing behavior (Buddington and Kuz’mina, 2000). Theseabass oral cavity (Dicentrarchus labrax) wasanalyzed in our laboratories by scanning electron

microscopy and light microscopy (Abbate et al., 2012).In this study the gilthead seabream (Sparus aurata,Linnaeus, 1758) was chosen because it is considered,with the seabass, the most diffused species for aquacul-ture in the area of the Mediterranean Sea, and as adirect consequence, the greatest share of the Mediterra-nean hatchery output is formed by these two species(Moretti et al., 1999). The gilthead seabream is a bonyfish belonging to the order Perciformes, family Sparidaeand it can be found in both marine and brackish waterenvironments such as coastal lagoons and estuarineareas, particularly during the early stages of its lifecycle. The gilthead seabream is mainly carnivorous,accessorily herbivorous and feed on shellfish, includingmussels and oysters (Bauchot and Hureau, 1990). S.aurata has traditionally been cultured in Mediterra-nean coastal lagoons and brackish/salt water ponds. Incaptivity, it is common in the Mediterranean Sea and ispresent along the Eastern Atlantic coasts, depth range1–150 m (Muus and Nielsen, 1999), usually 1–30 m(Lloris, 2005). Considering that the feeding habits incaptivity are one of the critical steps in the species ofcommercial interest, both during the larval phase and

F.A. and M.C.G. contributed equally to this work.

*Correspondence to: Francesco Abbate, Department of Morphology,Physiology, Biochemistry and Animal Productions, Faculty of VeterinaryMedicine, University of Messina, Messina, Italy. E-mail: abbatef@unime.it

Received 22 June 2012; accepted in revised form 17 July 2012

Contract grant sponsor: The University of Messina; Contract grant number:PRA 2008/2009

DOI 10.1002/jemt.22114

Published online inWiley Online Library (wileyonlinelibrary.com).

VVC 2012 WILEY PERIODICALS, INC.

MICROSCOPY RESEARCH AND TECHNIQUE 00:000–000 (2012)

the fattening, and that this process is strictly related tothe morphological characteristics of the oropharyngealcavity, this study was undertaken using light micros-copy and scanning electron microscopy to analyze themorphology of the dorsal tongue surface of the giltheadseabream to show if the anatomical characteristics ofthe tongue and the nutritional habits and choices ofthis farmed species can be closely related.

MATERIAL AND METHODS

Ten subjects of gilthead seabream (weight 5 250 gand length 5 15 cm) from the C.I.S.S. (Sicilian Centerof Experimental Ichthyopathology) of the University ofMessina, were used for this study. The fishes wereanesthetized with MS-222 (ethil-m-aminobenzoate, 0.2g/L) for 10 min and killed by decapitation. After thewithdrawal of the heads, for a good exposure of thetongue, the temporo-mandibular joints were disarticu-lated and cut. The samples were fixed in 2.5% glutaral-dehyde in Sorensen phosphate buffer 0.1M. Afterseveral rinsing in the same phosphate buffer, theywere dehydrated in a graded alcohols series, critical-point dried in a Balzers CPD 030, sputter coated with 3nm gold in a Balzers BAL-TEC SCD 050 and examinedunder a Cambridge Stereoscan 240 electron microscope(Zeiss ex Cambridge Instruments, Cambridge, UK)operating with an accelerating voltage of 20 kV. Forascertaining the morphological details better, somepieces were washed in 5% neutral Extran (Merck,Damstadt, Germany) to remove the mucus (Abbateet al., 2006, 2008, 2009, 2010, 2012). For light micros-copy the routine methods for hematoxylin & eosin andMasson trichrome staining were carried out.

RESULTSGross Anatomy

The tongue of S. aurata is not only a simple thicken-ing of the oral cavity floor, as often described in the

more diffused comparative anatomy textbooks. Themandibular branch and the skeleton branchial arcsform a skeletal scaffold to support the soft parts, whichform the floor of the oral cavity. This latter is particu-larly thickened to form a tongue with a triangularshape in which it is possible to identify an apex, a body,a root, and two lateral margins. The mucosa coveringthe dorsal surface of the body and the root is raised infolds with a lateral-medial trend, caudal-cranially con-verging on a median line in a raised ridge running thefull tongue, from the apex to the root. The dorsal sur-face of the tongue is widely irregular. The apex of thetongue is, with the mouth opened, completely free, con-versely, with the mouth closed, is enveloped in a sort ofpouch, with a Y shape, placed among numerous molari-form teeth arranged in 3–5 sets on each side to com-pletely cover the lower jaw (Figs. 1a and 1b).

Scanning Electron Microscopy

At the scanning electron microscope the surface ofthe pouch, enveloping the apical part of the tongue,showed the presence of foliate-like papillae on the sur-face of its apical and lateral part (Figs. 2a and 2b).Many mucosal plicae, longitudinally and parallellyorganized, characterize the main part of the pouch.Along the whole tongue surface, from the apex to theroot, an evident ridge is present: on the medial and lat-eral part of the apex many cylindroid papillae are pres-ent. The whole tongue is characterized by the presenceof two types of papillae, respectively with a fungiformand cylindroid aspect. They both are randomly distrib-uted through the whole dorsal surface of the tongue.The fungiform papillae, with an evident irregular sur-face (Fig. 3a), have a mechanic function. The cylindroidpapillae clearly protrude on the surface (Fig. 3c), sodemonstrating their mechanic role too. Scattered andnumerous taste buds were observed on the whole dor-

Fig. 1. Gross anatomy of the gilthead seabream oral cavity floor: Adrawing with different colors showing the tongue in green, the pouchin red, the molar teeth in yellow and the incisors in orange (a) and astereomicrograph (b) where the mucosal folds (arrows), the molar

teeth (arrowheads) and the incisors (white arrowheads) are evident.Scale bar: (b) 1 cm. [Color figure can be viewed in the online issue,which is available at wileyonlinelibrary.com.]

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sal surface of the tongue (Figs. 4a and 4b). Small andnumerous taste pores are also present, close to thetaste buds and distributed throughout the tongue sur-face (Fig. 5a).

Light Microscopy

By means of light microscopy it can be observed thatthe tongue is formed by an enveloping mucosa, a mus-culature, an osteo-fibrous skeleton. The tunica mucosa,at the dorsal surface of the tongue body is thickenedand is adhering to the underlying connective tissue.Some papillae with different size and shape, covered bya stratified pavimentous epithelium, are evident;particularly two different types of papillae, fungiform(Fig. 3b) and cylindroid (Fig. 3d), were observed on thewhole tongue dorsal surface. The connective tissue,raising beneath the epithelium, form the support to thepapillae. On the stratified epithelium, scattered tastebuds with the typical pear-onion shape and with theirapical part emerging from the epithelium surface, oftenassociated with taste pores, were observed not only onthe tongue surface (Figs. 5b and 5c), but also on thepouch (data not shown).

DISCUSSION

As previously described in other studies about fishoral cavity morphology (Abbate et al, 20062012), a veryimportant morphological finding of our study is thepresence of a true tongue. Up to now, a tongue-like wasdescribed in many teleosts only as a thickening of thepavement mucosa with a stratified epithelium (Bud-dington and Kuz’mina, 2000). The main characteristicof the gilthead seabream oral cavity floor is the pres-ence of an apical pouch, with, probably, a protectiverole mainly for the apical, free part of the tongue.

Moreover three zones, an apex, a body and a root canbe clearly distinguished, like in other teleosts (Abbateet al., 2006, 2012; Buddington and Kuz’mina, 2000).Our previous studies also described the presence in thezebrafish (Danio rerio) of a tongue formed by a medio-sagittal cylinder shaped structure, clearly isolatedfrom the jaw and adhered by differentiated folds to thelateral walls (Abbate et al., 2006). The tongue has afundamental role in the mechanics of food ingestionand, as described in the carp Cirrhinus mrigala byYashpal et al. (2009), the significant modifications inthe surface architecture at different regions of the oralcavity could be considered the adaptation related to thefeeding habits of the fish. These data are completelyconfirmed by our previous studies on the seabasstongue anatomy. As aforementioned, the gilthead seab-ream mainly eats shellfish, including mussels (Bauchotand Hureau, 1990), clearly influencing the morphologyof the tongue dorsal surface with particular regard tothe presence of numerous taste buds demonstratingthe interaction of food processing and taste, and there-fore the eventual reject of the nourishment. In S. aur-ata, like previously demonstrated in Dicentrarchuslabrax, an abundance of taste buds in the tongue indi-cates that this part of the oral cavity has specificallyevolved a peculiar gustatory system (Abbate et al.,2012) This could be highly responsive in evaluating thespecific food items to be consumed according to thepreference of the fish (Yashpal et al., 2009). It is wellknown that in fish the taste is strictly related to feed-ing and permits the discrimination among a variety offoods in an aquatic environment, giving the fishes thepossibility to distinguish the available to them (Hara,1994) and apparently playing an indispensable role inthe ultimate acceptance (Kubitza and Lovshin, 1997)

Fig. 2. The pouch enveloping a part of the tongue apex. (a) Foliate-like papillae can be observedthrough the whole surface of the apical and medial part of the pouch. (b) At higher magnification theinset of (a) with numerous foliate-like papillae. Scale bar: (a) 1 mm and (b) 250 lm. [Color figure can beviewed in the online issue, which is available at wileyonlinelibrary.com.]

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Fig. 3. Two types of papillae scattered on the whole tongue dorsalsurface. (a) A fungiform papilla with an irregular surface and (c) acylindroid papilla, both clearly protruding on the tongue surface.Scale bar: (a, c) 50 lm. (b) Light microscopy picture of a fungiform

papilla with the presence of taste pores (arrows) and (d) a cylindroidpapilla. 340. [Color figure can be viewed in the online issue, which isavailable at wileyonlinelibrary.com.]

Fig. 4. (a) A part of the tongue root showing a fungiform papilla with the presence of numerous tastebuds all around and close to it (arrows). (b) Inset of Fig. 4a at higher magnification with taste buds(arrows) scattered on the tongue root surface. (c) A particular of a single taste bud just beneath the epi-thelium. Scale bar: (a) 50 lm; (b) 25 lm; and (c) 3 lm.

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or rejection (Kruse and Stone, 1984) of potential fooditems. Many pores were also observed throughout thedorsal surface of the tongue and a chemosensory role,especially a taste role, could be hypothesized for them.The mechanical properties of the tongue are performedeither by the foliate papillae on the pouch, or the cylin-droid and fungiform papillae scattered on the tonguesurface. The cylindroid papillae, often oroaborallyoriented, have an aspect and, probably, a function ofprehension and chewing, a little bit similar to the sea-bass teeth present on the tongue surface. Our results,taken together, demonstrate that in the gilthead seab-ream important mechanic and sensory roles are carriedout by specific anatomical structures. Marked differen-ces are present between the seabass (Abbate et al.,2012) and the gilthead seabream tongue dorsal surface.The showed anatomical structures could be highlyresponsive in evaluating the specific food items to beconsumed according to the preference of the fish(Yashpal et al., 2009). In the two species aforemen-tioned the differences among the anatomo-functionalmechanisms related to the taste probably are in closerelationship with different level of perception. There-fore, a correlation is present between the food habits

and the need of a differentiation in the feeding man-agement of the seabass and the gilthead seabream, twospecies considered, up to now, very similar, oftenfarmed simultaneously using various types of ongrow-ing systems and fed following the same nutrition.Nevertheless, seabass and seabream are different spe-cies belonging to different taxonomic families and alsotheir susceptibility to stress and behavioral and physi-ological responses to stressful situations are known tobe different (EFSA, 2008). So with respect not only tothe commercial interests, but also to the animal wel-fare, further biochemical and physiological studies arenecessary to better know the differences in the me-chanical and sensory anatomo-functional aspectsbetween the gilthead seabream and the seabass, givingto our anatomical data an important support and con-firmation with the aim of improving the nutrition ofthese two species.

ACKNOWLEDGMENT

The authors thank Mr. V. Sidoti for his technicalassistance particularly for working on the electronmicrographs.

Fig. 5. (a) A taste pore (inside the blue inset) and a taste bud (inside the red inset). Scale bar: (a) 10lm. (b) Light microscopy pictures with the same morphological structures, the taste pore and (c) thetaste bud. 340. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.]

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