Nematoplana rubra, Curini-Galletti & Oggiano & Casu, 2002

Curini-Galletti, M., Oggiano, G. & Casu, M., 2002, The genus Nematoplana Meixner, 1938 (Platyhelminthes: Unguiphora) in eastern Australia, Journal of Natural History 36 (9), pp. 1023-1046 : 1024-1033

Nematoplana rubra sp. nov.

(gures 1, 4B, 8A, B, H; table 1) Synonym: Nematoplana sp. in Littlewood et al., 2000.

Material examined

HOLOTYPE: Australia, New South Wales, CoOEs Harbour, Arrawarra Headland , intertidal in medium sand (September 1993), whole mount (lactophenol): G211798 . PARATYPES: same data as holotype, three whole mounts (lactophenol): G211799– 211801.

Other material. New South Wales: two specimens from the type locality studied karyologically; CoOEs Harbour, Jetty, high to mid-intertidal in ne to medium sand (September 1993), one specimen studied karyologically; Port Stephens , Fingal Beach , mid-intertidal, medium to ne sand, three whole mounts (glycerine) (G211802 –4) (August 1996); Byron Bay , Tallow Beach , intertidal in medium sand, two karyological mounts (July 1993); Tweed River , outer beach, mid to lower intertidal in medium sand, four specimens sagittally sectioned (G211805 –8); two karyological mounts (August 1993) . Queensland: Coolangatta, Rainbow Beach , intertidal in medium to coarse sand: four specimens sagittally sectioned (G211809 –12), two whole mounts (glycerine) (G211813 –4), one karyological mount (August 1996); Kirra Point, ve specimens sagittally sectioned (G211815 –9); two whole mounts (glycerine) (G211820 –1); two karyological mounts; Peregian Beach, intertidal in medium sand (August 1993), one specimen .

Etymology The species is named after its distinctive coloration (latin ruber: red). Description

A medium-sized Nematoplana , torpid, fragile and easily broken during manipulations. The holotype, one of the largest specimens found, is about 4 mm long in xed condition. Animals crimson-red in life. Pigment apparently within parenchyma, particularly concentrated around brain and pharynx. Animals became colourless after xation in Bouin’s uid. Without pigmented eye-spots. Anterior end elongate, provided with few sensory bristles. Epithelium with non-depressed (intra-epithelial) nuclei, and entirely ciliated. Cilia about 3 Mm long. The distal edge of the epithelial cells is strongly eosinophilous. A thick basal lamina is present beneath the epithelium. Brain ovoidal, surrounded by a thin membrane. Subepidermal longitudinal musculature well developed on the ventral side.

With numerous spicules, arranged into a regularly spaced, dorso-median line. In the caudal region, spicules are often more numerous and appear less regularly arranged. Spicules (gures 1E, 8B) are about 10 Mm high, and consist of two umbrella-shaped hemispheres, joined by a straight shaft. The margin of the hemispheres is provided with numerous, sharp crenulations. The shaft is provided with a single spine, nearly at its middle. Spicules are dissolved by lactophenol (see below, in ‘Remarks’), and have been preserved in a few glycerine mounts only. Likewise, they have only been preserved in histological material xed by buOEered Bouin, with neutral pH. In sections, each spicule appears included within a cell. Spiculecontaining cells are usually found in the dorsal parenchyma, in some cases close to the basal lamina (gure 4B). Mostly, they show numerous, small granules of eosinophilous pigment, and appear glandular in nature. Cells closer to the basal lamina often lacked the eosinophilous granules. In some instances, spicule-containing cells have been found close to the gut: in one case at least, a spicule appeared to have been extruded from the pharynx, together with some gut content.

The gut extends posteriorly nearly to the caudal tip of the body, and, anteriorly, to just behind the brain. The short, collar-shaped, pharynx (gure 4B) is located in the posterior fourth of the body. It is ventrally orientated in mature specimens; in several sectioned immature specimens the pharynx was longitudinally orientated. The epithelium of the pharynx has depressed nuclei, and is ciliated except for a small area at the distal tip. Pharyngeal cilia are about 2 Mm long. Neither the oesophageal area nor the pharyngeal glands were observed. With weak longitudinal external and stronger inner circular musculature, particularly thick at the luminal side of the pharynx. Its basis is provided with a septum, which appears cellular in nature. The epithelium of the pharyngeal cavity is unciliated, with non-depressed nuclei.

Male genital organs. Very numerous testes in two rows among vitellaria, in front of the pharynx. The copulatory organ consists of two partly fused seminal vesicles, and a bulb provided distally with a stylet. The two seminal vesicles enter the bulb at its proximal base. They are lined by a thin epithelium. They are very elongate (about 500 Mm long in the holotype), and fused in their posterior half. Spermiducts enter the seminal vesicles anteriorly. Sperm are elongate, with long, spirally coiled nuclei. The bulb is ovoid-elongat e (about 55 Mm high in the holotype). It is lined with a coating of circular musculature, and provided with numerous prostatic glands. The stylet is a tubular structure, with a broad straight base, about 41 Mm wide in the holotype, and a narrow distal part, curving at about 180ss. The distal part is almost of the same diameter for all of its length. The base is strongly thickened at its ventral side, thus eOEectively reducing the size of the proximal opening to about three-quarters of the width of the base. A few longitudinal muscles, connected proximally to the bulb musculature, are attached to this thickened area, which thus at least functionally corresponds to an apophysis. The distal part of the stylet protrudes into a narrow, unciliated male antrum. The antrum opens to the outside through a male pore, which is located close to the mouth.

Female genital organs. The only two specimens which showed degrees of female maturity, radically diOEered in the morphology of the ovaries. The holotype (a specimen in full male maturity) had only two large oocytes, medially in front of the pharynx. The other specimen (which showed only traces of male maturity, being without seminal vesicles and with the copulatory bulb still in stages of formation) had instead two rows of ve isolated oocytes, laterally in front of the pharynx. Vitellaria stretch from behind the brain to the level of the female pore. The oviducts fuse at the level of the copulatory bulb into a common female duct, which opens to the outside through a female pore, surrounded by eosinophilous female glands, medially between the seminal vesicles.

Karyotype. With six chromosomes in its haploid set. Chroms 1–4 are metacentric, and distinctly larger than Chroms 5 and 6, which are more heterobrachial. Chrom. 6, in particular, is at the border between submeta- and subtelocentric. No obvious karyometrical diOEerence was apparent among the four populations sampled (gure 1G, 8H, table 1).

Discussion

The numerous, striking autapomorphies of the new species make discrimination from congenerics immediate: among the most obvious, no other Nematoplana is red, or contains spicules. The very large, recurved, base-less stylet is also distinctive. Only N. asita Marcus, 1950 has a somewhat comparable stylet, although much more swollen proximally, and spike-like distally (cf. Marcus, 1950). The stylet of N. rubra more closely resembles that of species of the genus Ezoplana Tajika, 1982 , which is however angled distally, and provided with a distinct apophysis (cf. Tajika, 1982). The presence of a cellular septum at the base of the pharynx, and the lack of pharyngeal glands, are further autapomorphic features of the new species.

The ovarian cycle of the new species appears peculiar, and suggests a diOEerential timing of male and female peak maturity. Data, however, are too scanty to allow any further inference.

Remarks

Based on similarity of pigmentation with other interstitial and entosymbiotic atworms ( Jennings and Hick, 1990; Crompton and Smith, 1963; Young and Harris, 1973), the red pigment is assumed to be globinic in nature. Globines are generally assumed to confer a competitive edge to species occurring in habitats with low P

( Jennings and Cannon, 1987). There is no evidence that N. rubra may occur particu- O 2 larly deep in sediment. However, where the species is common (generally, in the southern part of its known range), it is usually found in samples taken among large brown algae, stranded at low tide. Globines may thus enable the species to thrive in sediments reduced by microbial activity, in the presence of an overlying layer of decomposing organic material.

The spicules of N. rubra are particularly puzzling. Their ne morphology and size are strikingly identical, to the smallest details, to microscleres of Spongillidae (Porifera) (cf. Penney and Racek, 1968). That they dissolve in lactophenol, 5% acetic acid (as in karyological mounts), and non-buOEered Bouin points to a calcareous nature. They are preserved by alcohol, and in some of the glycerine mounts. In the latter case, diOEerential acidity of slides from diOEerent laboratories may be involved. However, spongillid microscleres are siliceous, and should be resilient even to the attack of strong acids. This notwithstanding, whole mounts in lactophenol and glycerine of microscleres of Spongilla lacustris (Linné, 1758) showed, after about one year, a more or less marked degeneration of shape (personal observation).

As understood by the authors, spicules are taken externally, presumably with food. They are then engulfed by apparently glandular cells, which migrate to the dorsal parenchyma, where they are arranged medially in a regularly spaced row, in a pattern closely reminiscent of the arrangement of cnidosacs (containing hydrozoan cnidae) of other nematoplanids and of species of Archimonocelis Meixner, 1938 (Tajika, 1982; Martens and Curini-Galletti, 1993). However, in no case were seen gut-pockets, such as those underlying cnidosacs ( Karling, 1966). Rather, spiculecontaining cells lie in the parenchyma, and a diOEerent functional pathway of transportation (see Karling, 1966) could thus be involved. Furthermore, apparently glandular cells such as those seen in N. rubra do not seem involved in the genesis of cnidosacs ( Karling, 1966). The activity of this type of cell may somehow etch Table 1. Nematoplana rubra sp. nov.: karyometrical data (number of plates analysed between the spicules, and make them more sensitive to a further attack by weak acids, as those used during specimen processing.

A further complication, however, arises from the fact that spongillids are only known from fresh-water habitats, and samples containing N. rubra were taken far from any obvious fresh-water in ux. However, marine diversity of Australia is still far from being exhaustively known, and the area may yield a still undetected marine spongillid, possibly epibiotic on the large brown algae around which, when stranded, N. rubra appears to be frequent. A simpler explanation would require a non-sponge origin, and a calcareous nature. However, the authors know no calcareous spicules similar in size and shape to those found in N. rubra . Holothuriid anchors, which are only marginally similar, are much larger, at and their extremities are never identical ( Frizzell et al., 1966).

Platyhelminthes are known to produce calcareous spicules. However, they are larger, extracellular structures, embedded in the parenchyma, with diOEerent, much simpler morphology. They are arranged all over the body, often in regular layers, and constitute a veritable ‘internal skeleton’ ( Rieger and Sterrer, 1975). The ‘cleptospicules’ of N. rubra may however perform a similar function, contributing to the rigidity of the body axis in a species lacking a chorda intestinalis.