SHELL AND BODY STRUCTURE OF THE PLESIOMORPHIC PULMONATE MARINE LIMPET SIPHONARIA PECTINATA (LINNAEUS, 1758) FROM PORTUGAL (GASTROPODA: HETEROBRANCHIA: SIPHONARIIDAE)

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INTRODUCTION
The siphonariids are marine pulmonate limpets with a combination of ancestral and derived characters (HubendicK 1946).They are herbivores (yonGe 1952), mainly inhabiting rocky shores of tropical and warm latitudes (dayRat et al. 2014).The detailed knowledge of their structure is crucial for the understanding of the evolution of Heterobranchia, the most heterogeneous group of gastropods, and the link between the opisthobranchs and the pulmonates.
The pulmonates are adapted to air breathing, using a pallial cavity modified into a "lung", connected to the exterior by an orifice called pneumostome and closed by a sphincter.Though most of them are entirely terrestrial, some primitive taxa are marine, and depend on the pelagic environment to complete their development.Within the family Siphonariidae, most taxa have planktotrophic veliger larvae (JabLonSKi & Lutz 1983, GRaHame & bRancH 1985), although some species undergo intra-capsular larval stage -a mode of development sometimes called direct development (PaL & HodGSon 2002), a term that has been used for the siphonariids (cHambeRS & mcQuaid 1994a, b, PaL & HodGSon 2003, 2005).Planktonic larval development is regarded as plesiomorphic within the family, and used as a systematic criterion to resolve their higher taxonomy and evolutionary relationships.The prevalence of this developmental mode supports the hypothesis that the siphonariids have a marine ancestry, rather than being derived from a terrestrial group which re-invaded the marine environment (HodGSon 1999 and references therein).
The genus and species concepts within the Siphonariidae have been challenged as a result of the use of non-conchological, i.e., phenotypic and molecular, data (e.g., dayRat et al. 2011, 2014, wHite & dayRat 2012).Forms of different shell structure have been proved to be conspecific variations, while conchologically similar forms have turned out to be different endemic species (e.g., HubendicK 1955).The anatomical information available is too tenuous to provide a strong support for a classifiction (e.g., HubendicK 1946); it leads to raising not easily recognisable or non-monophyletic taxa.There is a need for description of internal anatomical characters that could help delineating siphonariid species (dayRat et al. 2014).Currently, the research aiming at solution of the above mentioned problems focuses on molecular aspects rather than on anatomy.The relationships of the Siphonariidae, and of the Basommatophora in general, with the other Pulmonata, and their phylogenetic status within the clade Euthyneura, are also debateable; some molecular approaches indicate a closer relationship with the Sacoglossa opisthobranchs (GRande et al. 2004, 2008, dayRat et al. 2011, dinaPoLi et al. 2011, JenSen 2011).
Siphonaria pectinata (Linnaeus, 1758) has long been regarded as a north amphi-Atlantic species, occurring along the Atlantic coasts of Europe and North America (VoSS 1959).However, molecular analysis of samples from both sides of the North Atlantic suggests that these regions may hold two different species (KawaucHi & GiRibet 2011).Nevertheless, the species in its original sense has one of the broadest distributions in the genus, inhabiting the Mediterranean and the Eastern Atlantic Ocean from Portugal to Cameroon, with a gap at the Gulf of Guine (wHite et al. 2011).There are some indications that the range of S. pectinata in the Northeast Atlantic might be extending, but it is common only as far north as the Iberian Peninsula (S.J. HawKinS, pers. comm.).
Despite its importance, the morphological and anatomical knowledge of the siphonariids is relatively poor in view of their diversity and their nearly world-wide distribution.According to current estimates the genus Siphonaria comprises from 41 to over 100 recognised species (dayRat et al. 2014, WoRMS 2017).They are crucial for the understanding of the relationships of Pulmonata with the remaining higher heterobranchs.Inferring phylogeny within the pulmonates based on (mainly external) morphological characters is problematic due to the high incidence of homoplasy (wHite et al. 2011), and a more complete, holistic anatomical description can certainly throw new light on the question.Besides, despite their remarkable resilience to habitat disturbance (HodGSon 1999), the siphonariids are still much less studied ecologically compared to other limpets (e.g., patellogastropods), even in Europe, where S. pectinata is the only species present.Therefore, information on its anatomy may be crucial for understanding of several aspects of the species' ecology and its possible adaptations to environmental changes.The main objective of this paper is to describe and compare the morpho-anatomical features of a common and widespread siphonariid species, in order to provide a baseline for complementary studies on comparative biology, and possibly on phylogeny and ecology.

MATERIAL AND METHODS
The samples included large animals (shell length ~30-40 mm) collected on the rocky shores of the region of Sines, Portugal (Praia da Oliveirinha: 37°53'19.21"N,08°47'49.29"Wand adjacent areas), preserved in 70% ethanol, and deposited in the malacological collection of the Museu de Zoologia da Universidade de São Paulo (MZSP).The specimens were extracted from their shells and dissected using the standard techniques, under a stereo-microscope, with the specimen immersed in ethanol.Digital photos of most dissection steps were taken; camera lucida drawings were made; the drawings and description are based on all the examined specimens (N= 20).The radula was examined in scanning electron microscope (SEM) in the Laboratório de Microscopia Eletrônica of MZSP.Additionally, to illustrate some details of the external morphology, digital photos of two live specimens were taken in the field.

RESULTS
SIPHONARIA PECTINATA (LINNAEUS, 1758 (Figs 1-36) Shell 9).Limpet-shaped, up to 40 mm.Outline elliptical, width ~70% of length; height ~40% of length.Colour brownish grey to beige.Sculpture mosaic of ~100 narrow radial ribs, with rounded profile, gradually and uniformly increasing towards edge; interspaces slightly narrower than threads; concentric undulations and growth lines.Apex sub-central, slightly displaced to left and posterior (Figs 2-3).Inner surface dark brown, glossy, with narrow beige radial bands corresponding to external threads; brownish-beige spot in apical region occupying ~15% of surface.Edges slightly irregular; wide radial groove in middle of right-anterior quadrant (Figs 1-2); this groove marks a gap in the horseshoe-shaped muscle scars, located half way between apex and edge (Fig. 1).Apex eroded to various degree in all the studied specimens, from ~10% (Fig. 2) to almost entire dorsal surface (Fig. 9), as a result of environmental influence.For other details see VoSS (1959).
Head-foot (Figs 4,(6)(7)(8)20,22): of limpet shape, dorso-ventrally much flattened.Head uniformly yellow to cream-coloured with mosaic of greenish/ brown and creamy spots on its dorsal surface .Head in the form of thick anterior flap, anterior edge widely bi-lobed, medially grooved, each lobe moving relatively independent from the other; pair of minute eyes (Figs 6, 8: ey) immersed in integument, each one located half way between median line and head lateral edge.No tentacles or ommatophores.Ventral head surface transversely folded, with mouth (Fig. 6: mo) in middle.Foot (ft) wide and ample, flattened, occupying most of shell ventral surface; colour pale orange to grey; no appendages, except for wide flap below pneumostome (Fig. 4: fl), with ~1/5 of head-foot length, located at the level of shell groove.Shell muscle thick, horseshoe-shaped, surrounding almost entire shell edge, close to mantle edge (Figs 20,22: sm), except for anterior region (nuchal cavity) and right-anterior region (pneumostome) (both ~1/3 of shell width in length); left branch entire, right branch divided in anterior region, with ante-rior part of elliptical cross-section 22;im), and posterior region half as long as right branch.Dorsal surface of foot relatively plain, forming pallial floor (Fig. 20: pu).Pneumostome ventral flap (Figs 4, 6-8: fl) described below.Haemocoel occupying ~6% of head-foot volume, restricted by shell muscles to central regions of head-foot (Fig. 22) (for more details see below).
Radula , slightly longer than odontophore (Fig. 29), with rachidian teeth, plus ~35 (34-37, N=5) pairs of lateral teeth; no clear boundary with marginal teeth.Each radular row relatively straight in middle 2/3, marginal region slightly curved backwards (Figs 10,13,15).Rachidian tooth 14) small, ~0.2% of radular width, ~3 times longer than wide; base long and rectangular, cutting edge hook-like, elevating from posterior end, directed forwards; sharp pointed terminal cusp restricted, in dorsal view, to posterior third of tooth; pair of small but broad expansions at base of terminal cusp (Fig. 12).Lateral teeth similar to rachidian tooth 13), but twice as wide and cutting edge twice as long; form slightly asymmetrical, weakly bent internally.Cutting edge tip of lateral teeth varying from blunt  to bifid (Figs 14-18); in bifid cases, both terminal cusps similar, small, equidistant, turned forwards .Cutting edge of lateral teeth triangular, ~twice longer than wide; no basal cusp, but with longitudinal reinforcement as middle fold (Fig. 10  an).For other details, mainly histology, see KöHLeR (1894).

DISCUSSION
As in the case of most molluscs, the shell has for a long time been the main source of taxonomical knowledge on Siphonaria (HubendicK 1946).The shell of S. pectinata is characteristic and easy to identify due to its narrow and numerous radial ribs; despite this, it varies relatively widely.The variation (Figs 2,9) shows that erosion by waves can modify the shell surface and, to some degree, the shell height.However, the variation is comparable to that found in other siphonariids (e.g., tabLado & GaPPa 2001).The intertidal environment, with its highenergy water flow and wave action, influences the siphonariid shell shape and sculpture through some degree of phenotypic plasticity (cooKe 1911, teSKe et al. 2007).The phenotypic plasticity of the sipho-nariid shell shape and sculpture is regarded as ecologically adaptive in terms of intertidal zonation and geographical distribution: high-domed, light-coloured and more sculptured shells are found at higher shore levels and in tropical species (HodGSon 1999 and references therein); most of the samples analysed in this study came from high-energy sites.It is thought that some characters of the shield-shaped shells of Siphonaria can be influenced by the environment (wHite et al. 2011).
The siphonariids (for specific taxa see below) have some anatomical peculiarities that are so far exclusive; some of them are discussed below.Despite the limpet-like shell shape, the usual somatic modifications involved in the "normal limpetisation pro-cess" are not found in the siphonariids.The so-called "normal limpetisation process" will be dealt with in another paper, presenting the evolution from a typically coiled gastropod to an uncoiled, limpet-shaped one, in which all the structures are functionally restricted to the apertural region of the coiled snails.Most of the limpets examined within the project, for example patellogastropods (e.g., LeaL & Simone In those groups the visceral hump is conical, as an internal cast of the apical shell region.In fact, the placement of visceral structures in the head-foot haemocoel is normal in another process which is common among gastropods -the limacisation -in which a typically coiled snail becomes a crawling, shell-lacking head-foot.While in the so-called "true limpets" the respiratory cavity consists of a space between the mantle and the visceral mass opening to the external environment at its anterior end, the respiratory cavity of siphonariids, the "false limpets", is almost entirely closed, with a small contractile orifice on the right side, where the characteristic siphonal groove is located.The generic name Siphonaria was inspired by this furrow, certainly produced by the flap which protects the pneumostome . Another interesting difference between the siphonariid way of limpetisation and that of the remaining above-mentioned limpets is the shell muscle.In the so called "true limpets" it is horseshoe-shaped (in top view), with the muscle scar as an unbroken semiring on the internal side of the shell.Although the siphonariid shell muscle also looks like a horseshoe, its right branch is interrupted at about ¼ of its length 22: sm,im) in order to shelter the pneumostome (Figs 20,22: pn) and the anus (an); a corresponding interruption is present in the muscle scar on the inner surface of the shell.This conformation of the shell muscle is associated with the characteristically smaller tenacity (force of attachment to the substratum) and greater foot flexibility of siphonariids compared to the other limpets, and thus it has ecological implications, such as their restriction to habitats protected from direct wave action and their mobility as highly active grazers (HodGSon 1999 and references therein).It is easy to interpret the horseshoe-shaped shell muscle as derived from the columellar muscle of the coiled gastropods.This is probably the case for the siphonariids, as at least the main portion of the shell muscle may be homologous to the columellar muscle.However, its isolated portion located at the right-anterior end (Figs 20, 22: im) cannot be derived from the columellar muscle, since the genital structures surround it externally (Fig. 22: so).Thus there is no way to envisage modifying of the columellar muscle in order to derive the siphonariid organisation, with some haemocoelic structures contouring a part of the muscle externally.To our knowledge, no developmental studies have furnished a clue on this issue (e.g., Knox 1955).The best interpretation is that the main portion of the siphonariid shell muscle (sm) is actually derived from the columellar muscle; however, the isolated portion (im) may be a new acquisition, without any correlation with that of other gastropods.Unlike our observations of S. pecti nata, in S. tristensis ("Leach" in Sowerby I, 1823 = S. lessonii Blainville, 1827, see GüLLeR et al. 2016) the genital structures have been illustrated as running directly anteriorly, not surrounding the isolated branch of the shell muscle (im) (daLL 1870: pl. 5, fig.3); if this is confirmed, it will represent an additional siphonariid feature that merits further investigation.A possible misinterpretation is the shell muscle shape in S. gigas Sowerby I, 1825, which has been illustrated as continuous, only opening in the pneumostome area (HaLLeR 1893: figs 11, 12).On the other hand, and despite the above argumentation, the connection of the penis muscle with the isolated component of the shell muscle in S. pectinata (Fig. 22: in, pm) may be regarded as an indication that this portion of the shell muscle may be homologous to the columellar muscle.This contradictory issue certainly requires additional studies.In fact, the arrangement found in the heterobranch limpets has been mentioned by HaSzPRunaR (1985: 26) as being probably a result of the semi-detorted situation of the mantle/heart complex, also reflected by the euthyneuran nervous system.However, no conformation is found in the heterobranch ancylids (Simone et al. 2012).On the other hand, there is a strange similarity between the isolated component of the shell muscle (Fig. 22: pm) and the "adductor" muscle of some Sacoglossa, used for occluding the shell aperture by the flexible outer lip.This "adductor" muscle occurs, for example, in the genera Ascobulla Marcus, 1972 and Cylindrobulla Fischer, 1857, and is surrounded by some components of the genital ducts (maRcuS & maRcuS 1970; personal observation); in other words, it might be easier to explain the siphonariid limpetisation starting from a sacoglossan ancestor.Indeed, some molecular approaches have shown a close relationship between the siphonariids and the sacoglossans (GRande et al. 2004, 2008, dayRat et al. 2011, dinaPoLi et al. 2011, JenSen 2011).
The siphonariid head is also peculiar.It is a widely bi-lobed anterior flap (Figs 6-8: he), with a pair of eyes immersed in the integument (Figs 6-7: ey, 8) (daLL 1870, yonGe 1952).No other group of pulmonates has a similar head structure (tiLLieR et al. 1992, Simone 2011).Normally the head is convex and provided either with a pair of non-retractile lateral tentacles (basommatophorans) or two pairs of retractile ones (stylommatophorans).The eyes are normally associated with those structures, being located at the base of tentacles in basommatophorans, and on the tip of dorsal tentacles in stylommatophorans.The siphonariid model, in fact, resembles that found in some opisthobranchs, such as some doridaceans and their allies (e.g., aLVin et al. 2014), apart from the presence of rhinophores.
The dorso-lateral pedal defensive glands, epidermal multicellular structures not described here, appear to be an interesting characteristic of the siphonariids (PincHucK & HodGSon 2009).This kind of gland resembles that found in some calyptraeoidean caenogastropods (Simone 2002), although in that case the glands are distributed along the mantle border.In S. pectinata, an anti-predatory secretion is produced and exuded as a thick and white mucus (ocaña 2003), containing metabolites described as toxic (PauL et al. 1997) and in live specimens its lateral release is commonly observed when the animal is disturbed.Effective chemical defence against predation has been demonstrated for S. capensis Quoy et Gaimard, 1833in South Africa (mcQuaid et al. 1999), and is probably widespread in the genus, since at least 12 species have been found to biosynthesise polypropionates, compounds associated with the highly glandular nature of the foot-tissue and with their unpalatability (HodGSon 1999, mcQuaid et al. 1999, andreferences therein).A defensive response to predation is also present in intertidal pulmonate limpets of the allied family Trimusculidae, including release of a white secretion through the extended mantle, although in this case diterpenes have been identified as the predator deterring chemicals (San-maRtín et al. 2009).Though the ecological role of these secretions is mostly unknown, predation avoidance in an environment susceptible to both terrestrial and aquatic predators certainly has implications for several aspects of the snail's biology, such as longevity and foraging behaviour, and thus it may have an adaptive value for the siphonariids.
As air breathing pulmonates, the siphonariids have a well-developed "lung" (Fig 20: pu), in which the air is inhaled and exhaled though the pneumostome (pn) (yonGe 1952), and the flap ventral to it controls the breathing (Figs 6-7: fl, which is indicated in Figs 20, 22 as "pn"; "siphonal notch" by cottReLL 1910; "anal lobe" by maRcuS & maRcuS 1960).The normal pulmonate lung bears well-developed vessels, however this is not true of the siphonariids, which, instead, have a well-developed gill (Figs 5,19,21: gi), with a ctenidial vein running to the auricle (Fig. 21: cv).The location, structure and variation of the thick gill filaments suggest that the siphonariid gill is secondary, and non-homologous to the mono pectinate gill of other monotocardians (Simone 2011).This secondary condition of the siphonariid gill has long been accepted in the literature (ViLLieRS & HodGSon 1987).Besides, the siphonariid secondary gill appears to vary between species; those of S. pectinata (studied here) and S. hispida Hubendick, 1946(maRcuS & maRcuS 1960) appear to have a simpler structure than that of S. obliquata Sowerby I, 1825 (cottReLL 1910: fig. 1) and S. capensis (ViLLieRS & HodGSon 1987), in which the filaments have secondary folds; the gill of S. gigas is peculiar in being displaced to the periphery of the pallial cavity (HaLLeR 1893: figs 11, 12).The combination of gill and lung indicates that the animal is amphibious, breathing in both water and air (weLLS & wonG 1978, HodGSon 1999).Along with this ability, the respiratory physiology of siphonariids provides other advantages to cope with the harsh intertidal conditions, such as the facultative depression of aerobic metabolism in response to prolonged air exposure and desiccation; and the tolerance to hypoxic conditions and possible anaerobiosis on shores subjected to episodic sand inundation (HodGSon 1999).The anatomical basis for this physiological resilience is the withdrawal of the mantle skirt and the closure of the pneumostome, which close the connection of their vascular cavity with the external environment.These physiological adaptations enable the animal to survive and save energy in adverse habitats.As described above, the pneumostome of S. pectinata, and the siphonariids in general, lacks sphincter and cannot be closed (except by ventral flap).
The siphonariid reno-pericardial structures are relatively small for a limpet of such size, except for S. gigas, which has a relatively large kidney (HaLLeR 1893).The renal component touching the roof of the pallial cavity in S. pectinata (Fig. 19: ki) looks much smaller than the corresponding structures in its congeners (S. obliquata from New Zealand -cottReLL 1911; seven species of Siphonaria from South Africa -aLLanSon 1958; S. hispida -maRcuS & maRcuS 1960).However, the dimensions of the pericardium are similar across these taxa.The siphonariid digestive system also shows some interesting peculiarities.The odontophore is basically typical of most Pulmonata, bearing a well-developed pair of hard odontophore cartilages, widely fused with each other along the ventral-medial edge ; and the basic arrangement of intrinsic and extrinsic muscles.However, despite the relatively high degree of cartilage fusion, S. pectinata has a well-developed horizontal muscle , in which the anterior region overlaps with the cartilage fusion (Fig. 31).The pair of ventral tensor muscles of the radula (Figs 30-31: m11) is extraordinarily wide, even duplicated (m11, m11a); while in other pulmonates the pair of m11 is narrow and thin.This thick m11 conformation indicates that the radula is used in a way characteristic of hard-scraper snails.Besides the strong pair m11, most of the odontophore muscles are also relatively strong and thick (Fig. 20), showing an intense use of the radula, which has been also observed in other siphonariids (bLacK et al. 1988).Nonetheless, this contrasts with the shortening of the radula 28: rs).As a rule, the intense use of the radula results in its considerable elongation, as shown by littorinid caenogastropods (e.g., Simone 1998), in which the radula is longer than the shell.The most modified pair of odontophore muscles in S. pectinata is the m7 .The pair m7 connects the medial and middle edge of the pair m11a with the internal surface of the radular sac (Fig. 29: m7), crossing the middle level of the m6 .The function and homologies of the m7 pairs are enigmatic (Simone 2011), and in this respect S. pectinata is no exception.As the odontophore muscles of other siphonariids have not been subject to any other study, the interpretation of such differences is impossible, and they may be relevant at any taxonomic level.
The radular morphology of the siphonariids is typical of pulmonates.The radula is a carpet of small and relatively uniform teeth.In contrast to the normal pulmonate radula, the distinction between marginal and lateral teeth is difficult in most siphonariids, showing a gradual change from medial to lateral regions.In other siphonariids lateral and marginal teeth differ slightly, for example in Williamia radiata (Pease, 1860) in which the former teeth are strongly bicuspid, while the marginal ones are single plates (maRSHaLL 1981); this pattern is very different from the more homogeneous radula of S. pectinata.The radula of S. pectinata differs from that of S. hispida (maRcuS & maRcuS 1960: fig. 15) in having the rachidian still narrower, and in lacking multiple cusps at the cutting edge of the marginal teeth; it differs from that of S. obliquata (cottReLL 1910: fig.2) in having a narrower and sharper pointed rachidian; and it also differs from S. thersites Carpenter, 1864 and S. tristensis (actually S. lessonii) in having the sec-ondary cusps of the lateral and marginal teeth not so well-developed (daLL 1870: pl. 5, fig.1).It should be emphasised that in S. pectinata the radula can bear simple  or bifid (Figs 13-18) tips, with possibly variable excavating ability into the rocky substrate while foraging.Overall, the fine-toothed radula should be comparatively weaker than the large and strongly mineralised teeth typical of patellogastropod radulae (HodGSon 1999).These differences in the radular structure of microphagous grazers are probably translated to grazing capacity, influencing interactions among sympatric limpet taxa.In SW.Portugal, S. pectinata co-occurs with Patella depressa Pennant, 1777 and Patella ulyssiponensis Gmelin, 1791, the dominant limpet species of mid and low intertidal zone, respectively.Compared to these patellids, S. pectinata is generally much less abundant and its distribution is more variable in space (SeabRa in prep.).One of the possible mechanisms explaining this pattern might be an inferior competitive ability of S. pectinata in exploiting limited food resources (SeabRa in prep.).ocaña & Fa (2003) found that S. pectinata in Gilbraltar grazed exclusively on superficial soft algae, suggesting that this feeding specialisation could reduce competition with Patella limpets, which graze by deep scraping of the rocks and often include calcareous encrusting algae in their diet.In fact, siphonariids are often out-competed but not completely eliminated by patellogastropods (HodGSon 1999).
The siphonariid oesophagus is wide and has thick muscular walls (Figs 27,33: ec).This shows that the organ serves to store food, being a gizzard, and is capable of distension and additional mechanical food processing.The siphonariid stomach, on the other hand, is relatively small, being a single corner in which both ducts of the digestive gland lobes open (Fig. 27: st); the stomach of S. pectinata looks proportionally smaller than that of S. obliquata (cottReLL 1910) and of S. hispida (maRcuS & maRcuS 1960).The remaining digestive structures are typical of herbivore or microphagous snails, with several intestinal loops compressed by the remaining head-foot structures (Fig. 24: in).The formation of faecal pellets (Fig. 24: fp), instead of a single faecal string, is another interesting character in most siphonariids, as well as the location of the anus on the floor of the pallial cavity.In most gastropods which have pallial cavity the anus is located in its roof.The rectal region in S. gigas has a large bulging chamber (HaLLeR 1893: fig.16: ed), which apparently is unique in that species.
The simplicity of the intestinal loops in the siphonariids is noteworthy, compared to other herbivore limpets, being related to the fact that the digestive gland rather than the intestine performs most of the enzymatic and absorption functions (muRty et al. 2013).However, these functions of the digestive gland appear to be normal in Mollusca.In about half of the examined specimens the contents, apart from vegetal matter pulverised by the radula, included whole specimens of the minute caenogastropod Skeneopsis planorbis (Fabricius, 1780), and sometimes young specimens of the mussel Mytilus sp.These organisms were found in the gastric and intestinal contents, with the soft parts apparently intact.They might be merely contaminants, although it cannot be excluded that they serve as additional nutritive matter, which could explain the relative shortness of the intestine.
The genital system of S. pectinata has the same general structure as in the remaining siphonariids, having roughly the same main components (Hutton 1882).However, each known species appears to bear the following distinctive characters: S. pectinata (Fig. 32) differs from S. hispida (maRcuS & maRcuS 1960) in having proportionally smaller middle glands (albumen and capsule glands: ac, ag, which remain constant in specimens collected throughout the year) compared to the gonad; in the hermaphrodite duct (hd) being wider and blunter, and mainly in the structure of the copulatory organ, lacking flagellum and epiphallus, being a single bulk, while in S. hispida it is Y-shaped.The genital orifice of most siphonariids is mostly single and simple, located in the right-anterior region of the body edge, as described for S. pectinata (Fig. 22: gp).So far, the genital system of S. gigas is the only one with two orifices (HaLLeR 1893: fig 19).Preceding the genital aperture, there is in S. pectinata what used to be called penis (Fig. 32: pe), but actually it serves both female and male branches of the genital system.Inside (Fig. 35) it has a main, central, papilla-like component as outlets of the spermoviduct and of the bursa (ta), while the outlet of the penis gland is located posteriorly, outside the papilla (pd).This whole penis resembles those of the remaining pulmonates in being retractile, proboscis-like, and certainly its main body (Fig. 35: pe) may be everted during copulation, with the central papilla as the tip (ta, ap).The retraction of the structure is certainly provided by the penis retractor muscle (pm), which is very similar to those of the remaining pulmonates, mostly originating from columellar muscle.The penis muscle of the siphonariids originates from the isolated component of the shell muscle (Fig. 22: in,pm).It is interesting to note that protandry has been documented in S. pectinata (e.g., maRcuS & maRcuS 1960).
The siphonariid central nervous system is relatively uniform amongst the species, sharing some distinctive characters compared to other pulmonates, such as the wide distance between both cerebral ganglia (Fig. 36: ce); the asymmetry of the shape of the left and right pedal and pleural ganglia (pp); the proximity of the abdominal or parieto-supra-intestinal ganglia (zr); the different length of the connectives between the cerebral and the pleuro-pedal ganglion; and the particular conformation of the connective of the visceral (abdominal)-subintestinal ganglion (as).The nerve ring of S. pectinata is different from that of other siphonariids in having a single pair of pedal commissures, which is relatively thick, as it is also the case of Williamia gussoni (Costa, 1829) (RutHenSteineR 2006), while S. hispida (maRcuS &maRcuS 1960) andS. obliquata (cottReLL 1910) have this commissure thinner and duplicated.Another inter-specifically contrasting aspect is that S. pectinata and S. hispida (maRcuS & maRcuS 1960) have the pedal and pleural ganglia completely fused (PP), while they are close, but separated in S. obliquata (cottReLL 1910).The arrangement of connectives of the visceral (abdominal)-subintestinal ganglion (as) and the parieto-supra-intestinal ganglion (zr) appears to be slightly different in all species in which this feature was examined.
As mentioned above, living intertidally, the siphonariids are part of a complex ecosystem that highly influences their shell morphology.Their outer surface becomes a garden of epibionts, an example can be seen in Fig. 9 with the fouling of non-calcareous algae Ralfsia verrucosa (Areschoug, 1845).Besides, it is relatively common to find epibiotic fouling or filamentous algae covering the specimens living in more damp habitats, such as Caulacanthus ustulatus (Mertens ex Turner) Kützing, 1843 in the specimen in Figs 7-8.The colour of the head-foot also varies to some extent.This pertains mainly to the foot (rath-er than the head, which is usually yellowish) whose colour varies from yellowish orange to grey, possibly depending on the diet, habitat or the animal's size.Some of the spots are most probably related to the dorso-lateral pedal defensive glands (PincHucK & HodGSon 2009) which were not investigated here.
The phylogenetic placement of the family Siphonariidae is debateable, with supporters of all rational possibilities, from the traditional Basommatophora, to Archaeopulmonata, Opisthobrancha and Stylommatophora.For example, Siphonaria may be nested within opisthobranchs (GRande et al. 2008, wHite et al. 2011) or constitute the basal-most lineage of pulmonates (KLuSSmann-KoLb et al. 2008, dayRat et al. 2011).Of course, the contradictory results have been provided by molecular approaches, which in themselves are somewhat discordant and still under analysis.Concerning the morphological characters, any possible similarity to each of the above-mentioned taxonomic groups does not bear a closer examination.An example is the pallial cavity, whose resemblance across some of these groups has been interpreted as superficial, and explained as homoplasies in response to similar environmental pressure (JenSen 2011).
A discussion on the phylogenetic uniformity and the problems related to the siphonariids in particular, can be read in ScHRödL (2014).The issue will be further analysed in an ongoing paper (senior author).So far, at least from a morphological point of view (see: GiRibet 2015), some basic synapomorphies of Pulmonata can be evoked, which are present in both Basommatophora and Stylommatophora, such as the unique dorsal jaw plate; the retractile shape of penis; the penis retractor muscle originating from columellar muscle; the bursa copulatrix with elongated duct, inserting close to the genital orifice and placed close to the pericardium.Besides, though some siphonariids appear to have no penis, the general organisation of the genital system of the so far investigated siphonariids indicates an arrangement which is mostly similar to that found in as other plesiomorphic pulmonates (moRton 1955, RutHenSteineR & StocKeR 2009).
Though the phylogenetic relationships of the Siphonariidae are not within the scope of this paper, the facts and arguments briefly presented above should be at least considered.The problem will be dealt with in a future paper which will discuss representatives of several basal and higher pulmonates in a formal phylogenetic scenario.

Figs
Figs 31-34.Siphonaria pectinata.Anatomy: 31 -odontophore, ventral view, superficial membrane and muscles removed, right peribuccal muscle (mj, left in Fig.) removed; 32 -expanded genital system, dorsal view, topology of some adjacent structures and transverse section in indicated regions also shown; 33 -midgut, dorsal view, mostly opened longitudinally to show inner surface, topology of buccal mass shown; 34 -detail of middle region of genital system, dorsal view, showing insertion of some structures.Scale bars 2 mm
It differs from S capensis (aLLanSon 1958, PaL 2007) in having the hermaphrodite duct longer and more coiled, more divided in middle, and in a more elongated shape of the penis gland (pg).It differs from S. anneae Tomlin, 1944 (= S. carbo Hanley, 1858, see: teSKe et al. 2007) and from S. deflexa Born, 1778 (aLLanSon 1958) in having the penis retractor muscle (pm) much wider and in a more elongated shape of the penis gland (pg).It differs from S. obliquata (cottReLL 1910: fig.6) in having the hermaphrodite duct (hd) much thicker, the seminal receptacle (sr) much longer and wider, and the penis gland (pg) longer and coiled.It differs from S. virgulata Hedley, 1915 and from S. luzonica Reeve, 1856 in having the penis gland inserted in the posterior instead of the anterior region of penis (HubendicK 1955), the latter situation appearing to be the commonest (Hutton 1882).It differs from S. serrata (Waldheim, 1807) (PaL 2007) in having the penis gland inserted in the posterior end of the penis and not on its base, and in lacking flagellum in this gland.It differs from Williamia radiata (RutHenSteineR et al. 2007) in having a much smaller and single penis gland, inserted on the penis tip.For additional details of the siphonariid genital structures see PaL & HodGSon (2002, 2003, 2005) and PaL (2007).