Epidermal ultrastructure of the adult parasitic phase female and encapsulated larva of Kronborgia isopodicola (Platyhelminthes, Fecampiidae): Phylogenetic implications

lnrernnfional Journal/or Printed m Grear Britain

Parasirology

Vol. 23, No. 8, pp. 1027-1037. 1993 0

002&7519/93 IE6.00 + 0.00 Pergomon Press Ltd 1993 Ausrrolian Smielyfor Parasirology

EPIDERMAL ULTRASTRUCTURE OF THE ADULT PARASITIC PHASE FEMALE AND ENCAPSULATED LARVA OF KRONBORGIA ISOPODICOLA (PLATYHELMINTHES, FECAMPIIDAE): PHYLOGENETIC IMPLICATIONS J. B. WILLIAMS Department

of Zoology,

University

of Canterbury,

(Received I3 October 1992; accepted

Christchurch,

New Zealand

18May 1993)

AbStm2-WILLIAMS J. B. 1993. Epidermal ultrastructure of the adult parasitic phase female and encapsulated larva of Kronborgiu isopodicola (Platyhelminthes, Fecampiidae): phylogenetic implications. International Journalfor Parasitology 23: 1027-1037. The parasitic phase female K. isopodicola possesses a ciliated epidermis of polyhedral cells. Adjacent lateral plasma membranes are separated at intervals creating intercellular spaces. Epidermal cilia are anchored by a horizontal rootlet, opposite which a spur projects from the basal body, and a narrow vertical rootlet. The cytoplasm contains coated vesicles, and coated pits lie between microvilli. Large granular and vesicular bodies (rhabdoids) are scattered through the epidermal epithelium; in the epidermis of the encapsulated larva, granular rhabdoids are densely packed and slender, more compact bodies also occur. The compact, granular and vesicular bodies are probably morphological variants of the same epidermal structures, suggested to undergo sequential changes accomplished in later stages by lysosomal activity. Morphologically similar epidermal bodies are found in triclads. They are also characteristic of the parasitic genus Urustoma, which shares other ultrastructural features with K. isopodicola. The Neodermata may have arisen from parasitic turbellarian forms, at a more “primitive” level of organization than ancestors of the contemporary Rhabdocoela.

INDEX KEY WORDS: Platyhelminthes; Fecampiidae; Kronborgiu isopodicola; parasitic epidermal ultrastructure; cilia; microvilli; coated vesicles; rhabdoids; rhabdites; spermatozoa; phylogeny.

INTRODUCTION

THE Fecampiid turbellarians are internal, true parasites throughout the growth and maturation phase of their life history. In recent years, they have attracted attention as a possible sister-group of the Neodermata, the taxon comprising the major parasitic platyhelminth groups (see review and references cited by Rohde, 1990). Accordingly, an investigation was undertaken of a member of the genus Kronborgia Christensen & Kanneworff 1964, K. isopodicola from New Zealand waters. This recently discovered dioecious fecampiid infects isopods, and passes the nonparasitic reproductive phase in banana-shaped cocoons cemented to intertidal rocks. The species was described by Blair&Williams (1987), and an historical survey and discussion of fecampiid systematics was provided by Williams (1988a). Electron microscopical studies were concerned with spermatozoon ultrastructure and phylogenetic implications (Williams, 1988b), the oocyte (Williams, 1989) vitellocyte (Williams, 1990a), epidermis of the parasitic phase

turbellarians; platyhelminth

male (Williams, 1990b), subepidermal glands (Williams, 1990~) and genital system (Williams, 1990d) of the female, and some aspects of larval ultrastructure (Williams, 1991). Watson, Williams & Rohde (1992) investigated the ultrastructure of the differentiated and developing larval eye, and Watson, Rohde & Williams (1992) described the larval protonephridial system. Bresciani & Koie (1970) and Koie & Bresciani (19173) recorded the epidermal fine structure of the female and larval ultrastructure of a related species, K. amphipodicola, endoparasitic in amphipods. Light and electron microscopical studies of the mature parasitic phase male reveal an epidermis characterized by unusually large, plentiful dense bodies occupying a major portion of the epithelium (Williams, 1990b). Epidermal bodies of comparable size and abundance occur in a triclad (Tyler, 1976, 1984), the kalyptorhynch Polycystis naegelii (see Schockaert & Bedeni, 1977), and Urastoma (see Noury-Srai’ri, Justine & Euzet, 1990). 1027

1028

J.~.WIL~IAM~

FIG. I, parasitic female. Oblique tangential section through epidermis. Scale line, 2 grn. agd, Acidophilic gland ducts; Ci, cilia; hr, horizontal cifiary rootlet; is, intercellular space; Md, mucus gland duct; N, nucleus;No, nucleolus; sbd, duct of gland producing small secretory bodies; VB, vesicular bodies. Stars, lateral pIasma membranes.

Epidermis of K. isopodicola

1029

FIG. 2. Parasitic female. Oblique tangential section. Scale line, 2 pm. agd, Acidophilic gland duct; cgd, cyanophilic gland duct; is intercellular space; mvb, multivesicular body; N. nucleus; VB vesicular bodies. Stars, lateral plasma membranes. Arrow, vesicular body discharging into intercellular space.

Refractile rodlets are produced by parenchymal glands in the female (Blair&Williams, 1987; Williams, 1990~). The rodlets, strongly eosinophilic and passed through microtubule-supported cell extensions to the epidermal surface, are referred to as rhabdites in the present paper. Their formative stages are unknown.

were stained with uranyl examined and photographed microscope. Free swimming capsules, were examined live

OBSERVATIONS Epidermis

MATERIALS AND METHODS Mature females were dissected from their host, the marine isopod Exosphaeroma obtusum, collected at Kaikoura, South Island, New Zealand. They were fixed in 4% glutaraldehyde in 0.025 M phosphate buffer for 6 h at room temperature, washed in three changes of phosphate buffer, then post-fixed in 2% osmium tetroxide in phosphate buffer for 5 h at room temperature. Specimens were dehydrated in an acetone series, and embedded in Spurr’s epoxy resin. Ultrathin transverse, longitudinal and tangential sections were cut from the surface in the middle third of the body. Egg masses removed from cocoons prised from rocks at Kaikoura were prepared for electron microscopy by the same protocol, and sectioned with enclosed larvae randomly oriented. Sections

nitrate and lead citrate, and with a JEM-1200 EX electron larvae, newly hatched from egg by light microscopy.

The

of the parasitic

epidermal

phasefemale

epithelium

is composed

of multi-

in near-tangential sections (Figs. 1,2). Adjacent ceils are connected near the surface by junctional structures (Fig. 8, j). Lateral plasma membranes (Figs. 1, 2, *) are closely apposed except in particular domains where clearly defined intercellular spaces exist (Figs. 1,2, is). Epidermal cilia (Figs. 1,3,9, Ci; Fig. 4) are of the usual 9 + 2 type, the axial microtubules (Fig. 4, am) terminating above the basal plate (Fig. 4, bp) without the intervention of an axosome. Each cilium is anchored by a strongly developed, cross-striated principal (horizontal) rootlet (Figs. 1, 3-6, hr). A slender accessory (“vertical”) ciliated

cells

which

are

polygonal

J. B. WILLIAMS

FIG. 3. Parasitic

FIG. 4. Parasitic

female. L.S. epidermal

surface,

showing ciliary rootlets. Scale line, 0. I pm. Ci, Cilium; hr, horizontal Arrow vertical rootlet.

female. L.S. epidermal surface, showing ciliary microtubules; bp, basal plate; hr, horizontal

structure and rootlets. rootlet. Arrow, vertical

Scale line, 0.2 pm. rootlet.

rootlet.

am, Axial

FIG. 5. Parasitic female. L.S. epidermis, showing microvilli, ciliary rootlets and coated vesicles. Scale line, 0.2 pm. co, Coated intermicrovillar pit; cv, coated vesicle; f, filaments of microvillus; hr, horizontal rootlet. Arrow, vertical rootlet. FIG. 6. Parasitic

female. Near-tangential

section. Scale line, 0.2 pm. co, Coated spur.

rootlet with cross-striations is also present (Figs. 3-5, arrow), oriented at an acute angle to the horizontal rootlet. A small spur projects from the basal body opposite the horizontal rootlet (Fig. 6, sp); fine filaments radiate from the tip of the spur into the cytoplasm. The epidermal surface is bordered by microvilli (Fig. 8, mv; Fig. 5) containing parallel filaments (Fig. 5, f). Coated intermicrovillar pits (Figs. 5,6, co) open to the exterior; in addition to an external coat, the pits possess a thickened membrane of increased density and a filamentous content. Coated vesicles (Fig. 5, cv) are found in the cytoplasm. Large,

intermicrovillar

pit; hr, horizontal

rootlet;

sp,

dense cylindrical bodies oriented perpendicular to the surface occur at intervals throughout the epidermis (Figs. I, 2, 7-10). These structures are of two kinds: granular bodies (Fig. 7), and more numerous vesicular bodies (VB) (Figs. 2, 9, 10, VB). The former have a finely granular or flocculent composition, and typically possess a narrow lamina of high electron density immediately underlying the bounding membrane (Fig. 7). Tubular inpushings of the bounding membrane and vesicles associated with an expansion of the marginal dense material are seen to penetrate into the interior of the granular bodies (Fig. 7, *). VBs

Epidermis of K. isopodicola

FIGS. 7-10.

1032

J. B. WILLIAMS

contain tightly packed, membranous vesicles and vacuoles with a clear content. They are surrounded by a thick coat of dense material lying external to the limiting membrane (Figs. 9, 10). The coat consists of two layers, an inner homogeneous layer of moderate electron density, and an outer, strongly opaque layer composed of granules or minute peglike elements (Figs. 9, 10, arrow). Elongated mitochondria, containing prominent intramitochondrial granules (Fig. 8, ig), are oriented at right angles to the surface. Many mitochondria are closely interrelated with the VBs, protruding deeply into the coating material (Figs. 9, 10, M). Small clear vesicles (Fig. 9, ve), granular vesicles (Fig. 9, gv), bodies containing flocculent matter (Fig. 10) and multivesicular bodies (Fig. 2, mvb) also occur in close spatial association with VBs. In places VBs are observed to discharge empty-seeming membrane configurations into intercellular spaces (Fig. 2, arrow). GER is weakly represented and Golgi systems are small and only seldom encountered. Nuclei (Figs. 1,2, N) are intraepidermal, and basally located. Nearly all nuclear chromatin is finely dispersed; patches of heterochromatin are small and sparse, and there is no peripheral layer of condensed chromatin. Nucleoli (Fig. 1, No) are centrally placed. Ribosomes bind to the outer nuclear membrane. The epidermis rests on a fully developed basement membrane of the polyclad/neoophoran type, consisting of a dense basal lamina and a distinct “fibrous” layer (Fig. 11, fl). Cytoplasmic processes of the two major unicellular gland types of the female, cyanophilic gland ducts (Fig. 2, cgd) and ducts of acidophilic glands implicated in the production of cocoon materials (Figs. 1,2, agd), pass through the epidermis. Mucus-secreting cell ducts are plentiful in the epidermal epithelium (Fig. 1, Md), and many processes of mucus glands underlie the basement membrane (Fig. 11, Mp). Slender ducts of parenchymal glands producing small secretory bodies additionally traverse

the epithelium (Fig. 1, sbd; Fig. 10). Ducts of parenchymal glands as a rule pass through individual epidermal cells, enveloped by expansions of the cells’ plasma membranes (Fig. 10, me), and are supported by cylinders of microtubules (Fig. 7, mt; Fig. 10). Rhabdite gland ducts, both penetrating the epidermis (Fig. 8) and in subepidermal sites (Fig. 1 l), are supported by microtubule sheaths (Figs. 8, 11, mt). The rhabdites are unlamellated and, in comparison with VBs, are noticeably regular in shape and outline (Figs. 8, 11, R). Together with a subepidermal rhabdite-transporting duct and a mucus gland extension, Fig. 11 illustrates an acidophilic gland duct passing through a perforation in the basement membrane (Fig. 11, *), and a process of a gland synthesizing small secretary bodies (Fig. 11, sbd). Single uniciliate receptors of the nonrecessed type were observed, and a study of these sensory organelles is projected. Epidermis of the encapsulated larva The larval epidermis differs from that of the parasitic female, as the epidermal bodies are abundant and closely packed, accounting for the major part of the epithelial volume (Figs. 12, 13). Apical ends of epidermal bodies occupy much of the free surface, to such a degree that microvilli are unevenly distributed, tending to occur in tufts. Most epidermal bodies are of the granular kind, or take the form of narrow, more compact structures of high electron density (Fig. 13, eb); additionally, bodies intermediate in density and texture are found. VBs are present but infrequently seen. Vesiculated areas of the granular bodies discharge apparently empty membrane loops to the exterior space (Fig. 12, arrow), and free circular membrane configurations are observable (Fig. 12, *). Granular epidermal extrusions also overlie the surface (Fig. 12, ep). Figure 13 illustrates the organization of the epidermis directly above the larval eye, a light gathering

FIG. 7.Parasitic female. T.S. epidermis; granular body, parenchymal gland duct and granular vesicle. Scale line, 0.5 pm. gv, Granular vesicle; mt, microtubules. Star, vesicles and expansion of peripheral dense layer in interior of granular body. FIG. 8. Parasitic female. T.S. epidermis; rhabdite and rhabdoids. Scale line, 0.5 pm. ig, Intramitochondrial junctional structures; mt, microtubules; mv, microvilli, overlying rhabdoids; R, rhabdite in duct.

granules;

j,

FIG. 9. Parasitic female. T.S. epidermis, showing vesicular bodies. Scale line, 0.25 pm. Ci, Cilium; gv, granular vesicle; M, mitochondrion closely associated with vesicular body; VB, vesicular body; ve, small clear vesicles associated with vesicular body. Arrow, coating layer of granular material, external to homogeneous layer enveloping vesicular body. FIG. 10. Parasitic female. Tangential section. Scale line, 0.5 pm. is, Intercellular space; M, mitochondria closely associated with vesicular bodies; me, expansion of plasma membrane enveloping parenchymal gland duct containing small secretory bodies and peripheral microtubules; VB, vesicular body. Stars, lateral plasma membranes. Arrow, coating layer of granular material, external to homogeneous layer enveloping vesicular body. Top centre, bodies containing flocculent matter.

Epidermis

of K. isopodieola

1033

ec

FIG. 11. Parasitic female. T.S. Epidermal basement membrane and unicellular gland processes. Scale line, 2 pm. A, Fibrous layer of basement membrane; Mp, mucus gland process; mt, microtubules supporting rhabdite duct; R, rhabdite; sbd, small secretory bodies in process. Star, acidophilic gland duct. FIG. 12. Encapsulated larva. Oblique tangential section showing closely packed granular bodies. Scale line, 1 pm. ep, Epidermal extrusion (?). Star, discharged membrane. Arrow, granular body extruding membrane loops. FIG. 13. Encapsulated larva. T.S. epidermis above eye. Scale line, 2 pm. eb, Slender, highly opaque epidermal bodies; ec, egg capsule; mv, photosensitive microvilli; N, expelled nucleus; rp, reflective platelets overlying nucleus of mirror cell of eye; sbd, small secretory bodies in duct.

J. B. WILLIAMS

1034

organ of the reflector type consisting of a mirror cell containing reflective platelets (Fig. 13, rp), and photoreceptive neurons with dendritic processes ending in bundles of long, photosensitive microvilli (Fig. 13, mv). The epidermis overlying the eye is not clarified in the living animal, and displays no ultrastructural modification. Not only are the epidermal bodies represented as usual, but in addition, ducts conveying small secretory bodies from parenchymal glands pass through perforations in the basal lamina and traverse the epidermis in this region (Fig. 13, sbd), and appear to release their secretion into the intracapsular space. Also shown in Fig. 13 is a degenerating nucleus (Fig. 13, N) evidently expelled from the larva; the expelled nucleus contains dispersed chromatin and condensed chromatin masses, together with clumped material of high electron opacity. Egg capsules often show an irregular contour (Fig. 13, ec).

DISCUSSION Epidermal

ultrastructure

In the absence of an alimentary tract, the epidermis of the parasitic female K. isopodicola functions as a digestive epithelium, and the epidermal epithelium of the encapsulated larva may also serve as an absorptive tissue (Williams, 1991). It appears nutrient molecules of low molecular weight from the immediate environment may pass across the free surfaces of the epidermal cells (Williams, 1988b, 1991). Some adult females collected were closely wrapped around the gut of the host, and so were ideally situated to promptly utilize breakdown products of the host’s digestive processes. Horizontal rootlets of locomotory cilia of K. isopodicola are directed anteriorly (Williams, 1990b). The strongly developed horizontal rootlet, together with the vertical accessory rootlet, presumably provides anchorage during the effective stroke of ciliary action. The spur projecting from the basal body opposite the horizontal rootlet may assist to support the cilium during the relaxed recoil phase. Similar projections from ciliary basal bodies have been observed in other turbellarians (e.g. in the dalyellioid Didymorchis by Rohde, 1987; the umagillids Cleistogamia longicirrus and Seritia stichopi by Rohde, Watson & Cannon, 1988; the umagillid Syndisyrinx punicea by Rohde & Watson, 1988; and Urastoma cyprinae by Noury-SraYri et al., 1990, Figs. H, I). Coated intermicrovillar pits are formed from coated vesicles which fuse with the apical plasma membrane. It is proposed that the vesicles open to the exterior, are flattened, raised above the surface and everted to form the tips of microvilli (see also Williams, 1982).

Microvillar tips are interpreted as hemidesmosomes elevated above the general surface level. Filaments contained in a microvillus may be regarded as a bundle of tonofilaments. The significance of the nucleus presumably expelled from the larva, shown in Fig. 10, is obscure. The extruded nucleus appears pycnotic; however its chromatin is largely in the dispersed configuration, distinct from the highly condensed chromatin characterizing nuclei of many cell types during late intracapsular development (Watson et al. 1992; Williams, unpublished observations). Studied by light microscopy, rhabdites of the female K. isopodicola are refractile and strongly eosinophilic; by contrast the cylindrical epidermal bodies or rhabdoids, most readily observable in the mature male, stain strongly but are not predominently acidophilic or cyanophilic (Williams, 1990b). As mentioned earlier, the rhabdites are conveyed to the epidermal surface by way of microtubule-supported ducts. Although they are nonlamellated, they might nevertheless represent true rhabdites of the triclad type, which do not have a lamellar substructure (Smith, Tyler, Thomas & Rieger, 1982). In formative stages, developing rhabdites of the triclad Procotylajuviatilis are composed of a peripheral cross-striated lamella enclosing a lucid matrix containing dense granules. The granules coalesce and the differentiated secretory body is entirely electron opaque (Lentz, 1967). The smaller, homogeneously dense rhabdoids observed in the larval K. isopodicola are believed to undergo a constitutional change, giving rise to large, less opaque granular bodies. The granular bodies in turn evidently transform into VBs by a process involving progressive membranous vesiculation; interruptions of the bounding membrane of granular bodies by canalicular invaginations, accompanied by expansions of the associated peripheral dense lamina, lend support to this view and probably represent an early stage in the vesiculation process. As vesiculation advances, external coating material simultaneously appears in the adjacent cytoplasm. Vesicular epidermal bodies are morphologically reminiscent of lysosomes in vermiform stages of the mesozoan Dicyema, believed to digest both ingested and endogenous material (Ridley, 1968) and of lysosomes effecting intracellular digestion in the intestinal epithelium of the scutariellid Troglocaridicola mrazeki (see Williams, 1992). The epidermal bodies have been interpreted as sites of lysosomal activity (Williams, 1990b, 1991) in the final stages of which they are filled with tightly packed, empty-seeming vesicles and vacuoles eventually eliminated as closed membrane configurations. The mechanism of discharge resembles neither merocrine nor typical

Epidermis of K. isopodicola apocrine secretion. Some VBs void membrane loops into intercellular spaces; and the bodies are closely associated with mitochondria and multivesicular bodies. These are features characteristic of lysosomes (Williams, 1983, 1984, 1993). Nutrients from the environment or yolk in the larva, or wastes from the epidermis or other body tissues, may undergo demolition within the epidermal bodies. Alternatively, the epidermal rhabdoids may be secretory bodies undergoing constitutional and conformational changes influenced by lysosomal enzymes. Lysosomal hydrolase activities might possibly contribute to a normal secretory process, although this appears unlikely in view of the observed discharge of some VBs into intercellular spaces; or the epidermal bodies may be secretory organelles which undergo lysis by cellular autophagy when they become surplus to requirements or senescent. The epidermal cells contain little machinery for protein synthesis, as Golgi systems are small and sparse and few GER cisternae are found. The immediate impression gained from fine structural study of this cell type, particularly when comparison is made with rhabditogenic cells of other turbellarians, is that the cellular synthetic machinery is too weakly developed to be capable of manufacturing the large inclusion bodies. Nevertheless, the epidermal rhabdoids might represent secretory organelles synthesized in epidermal formative or replacement cells. Large, vesiculated or vacuolated epidermal bodies were discovered by Bresciani & K&e (1970) in the cocoon phase female of K. amphipodicola. Phylogenetic considerations In the vesicular condition, the epidermal bodies of K. isopodicolu are morphologically very similar to those of an undescribed triclad (Tyler, 1984), the eukalyptorhynch Polycystis naegelii (see Schockaert & Bedini, 1977) and the parasitic Urastoma cyprinae (see Noury-Srai’ri et al., 1990). Urastoma is presently grouped with the Prolecithophora, although its of the distinctive prospermatozoon is not lecithophoran type and this classification is in doubt (Ehlers, 1988). These rhabdoid structures probably fall into the general category of epidermal bodies of the triclad kind as defined by Smith et al. (1982) characterized as rhabdoids of large size (Tyler, 1984) with homogeneous, often dispersed, flocculent contents (Smith et al., 1982). Spermatozoa of K. isopodicolu (see Williams, 1988b), Urastoma (see Noury-Srai’ri, Justine & Euzet, 1989~) and the kalyptorhynchs (Hendelberg, 1977) possess incorporated axonemes and mitochondrial derivatives in the form of long rodlike structures. On available evidence Kronborgia, Urastoma and Poly-

1035

cystis therefore have similar epidermal and spermatozoon ultrastructure. Further features common to K. isopodicola and Urastoma cyprinae are as follows: a parasitic mode of life; both practise epidermal insemination, the spermatozoa migrating through the parenchyma to the female ducts; the spermatozoa have two elongate mitochondrial derivatives and lack the dense bodies present in the sperms of many turbellarians. It appears from these data that Kronborgia’s closest affinities may lie with Urustoma in particular, and with other turbellarians at a relatively “primitive” level of organization, rather than with the Dalyellioida/Temnocephaloidea. Dalyellioids and temnocephaloideans possess small secretory epitheliosomes (Bedeni & Papi, 1974; Williams, 1975, 1980a: Noury-Srai’ri, Justine & Euzet, 1989b), and spermatozoa with free flagella (Williams, 1983, 1984, 1986; Rohde, 1987) with the exception of Paravortex whose sperm have neither flagella nor incorporated axonemes (Cifrian, Garcia-Corrales & Martinez Al&, 1988; Noury-Sra’iri, Justine & Euzet, 1989a). The larval K. isopodicola possesses a protonephridial terminal organ with a poorly developed weir apparatus and with the flame cell nucleus located close to the base of the ciliary tuft (Watson, Rohde & Williams, 1992). Urastoma has an atypical weir-like structure, formed from extensions of both the nucleated terminal cell and the adjacent tubule cell; the processes coil around each other and are not strengthened by microtubules (Rohde, Noury-Srai’ri, Watson, Justine & Euzet, 1990). Among the aforementioned group of turbellarians the kalyptorhynchs are exceptional: their protonephridial end organ, as described by Rohde, Cannon & Watson (1988) is a flame bulb of the kind typical of Rhabdocoela (sensu Rohde, 1990, Fig. 9) characterized by a weir comprising one row of microtubule-supported rods contributed by a single cell. The corresponding nucleus is located in an unknown position remote from the flame. Spermatozooa of K. isopodicola and Urastoma have incorporated axonemes, and therefore possess the platyhelminth sperm morphology generally interpreted as the most “advanced” or modified; it is therefore puzzling that these spermatozoa reach the oocytes by the comparatively primitive method of migration through the parenchymal tissues, those of the fecampiid insinuating between closely apposed plasma membranes of adjacent cells (Williams, 1990a, d). In order to reach the oocyte, the spermatozoa must be capable of sustained active, directed locomotion through compact tissues. A contrasting condition characterizes the temnocephaloidean species Temnocephalu geonoma Williams, 1980b, which like other typical members of the taxon Rhabdocoela possesses

J. B. WILLIAMS

1036

spermatozoa with free flagella, the cytomorphology generally believed to be relatively primitive. In this species, the function of sperm transport is usurped by the ovo-vitelline duct; spermatozoa, wound into a ball, are passively transferred through the lumen to the oocytes (or seminal vesicles) by reverse peristaltic contractions of the duct walls. Rohde (1990, 1991) convincingly argues that the Neodermata may have arisen from turbellarian forms less modified than dalyellioids and temnocephaloideans, and believes the Fecampiidae is possibly the sister-group of the Neodermata (also Rohde et al., 1990; Watson, Rohde & Williams, 1992). Similarities between the Neodermata and Kronborgiu include a protonephridium with flame cell nucleus adjacent to the base of the ciliary tuft, and features common to the Neodermata and Uruszoma include a protonephridial end organ comprising process from two cells, the flame cell and the proximal tubule cell. None of the three groups is characterized by a protonephridial weir apparatus composed of a single series of microtubulesupported ribs arising from a flame bulb, as found in the Rhabdocoela. All possess spermatozoa with long mitochondrial derivatives and lacking dense bodies. With a few exceptions among the Neodermata, all have spermatozoa with incorporated axonemes which reach the oocytes by passage through the parenchyma or copulation canals as opposed to via the ovo-vitelline duct; and all are parasites. If future studies reinforce this view of neodermatan origins from turbellarians at a relatively primitive level of organization, it may be necessary to revise the current cladograms symbolizing platyhelminth phylogeny (for detailed review see Rohde, 1990). Acknowledgements-Grateful thanks are extended to Dr Nikki Watson of the Department of Zoology, University of New England, Armidale, N.S.W., Australia, for valuable assistance, and to Mr Manfred Ingerfeld of the Department of Botany, University of Canterbury, for technical aid. The author is also indebted to Miss Jan McKenzie of the Department of Zoology, University of Canterbury, for technical assistance, and to Mr Terry Williams of the same Department for help with photography. REFERENCES BEDENIC. & PAPI F. 1974. Fine structure of the turbellarian epidermis. In: Biology of the Turbellaria (Edited by RISER N. W. & MORSE M. P.), pp. 108-147. McGraw-Hill, New York. BLAIR D. & WILLIAMS J. B. 1987. A new fecampiid of the Kronborgiu (Platyhelminthes: Turbellaria: genus Neorhabdocoela) parasitic in the intertidal isopod E.uosphaeroma obfusum (Dana) from New Zealand. Journalqf’Natural History 21: 1155-l 172. BRESCIANIJ. & K~IE M. 1970. On the ultrastructure of the epidermis of the adult female of Kronborgia amphipodicola

1964 (Turbellaria, Christensen & Kanneworff, Neorhabdocoela). Ophelia 8: 209-230. CHRISTENSENA.M. & KANNEWORFF B. 1964. Kronborgia amphipodicolu gen. et sp. nav., a dioecious turbellarian parasitizing ampeliscid amphipods. Ophelia 1: 147-166. CIFRIAN B., GARCIA-C• RRALESP. & MARTINEZAL&. S. 1988. Ultrastructural study of spermatogenesis and mature spermatozoa of Puravortex cardii (Platyhelminthes, Dayellioida). Acra Zoologica 69: 195-204. EHLERS U. 1988. The Prolecithophora - a monophyletic taxon of the Platyhelminthes? Fortschritte der Zoologiel Progress in Zoology 36: 359-365. HENDELBERG J. 1977. Comparative morphology of turbellarian spermatozoa studied by electron microscopy. Acta Zoologica Fennica 154: 149-162. K~IE M. & BRESCIANIJ. 1973. On the ultrastructure of the larva of Kronborgia amphipodicola Christensen and Kanneworff. 1964 (Turbellaria, Neorhabdocoela). Opheliu 12: 171-203. LENTZ T.L. 1967. Rhabdite formation in planaria: the role of microtubules. Journal of Ultrastructure Research 17: I14126 NOURY-SRAIRI N., JUSIINE J.-L. & EUZE-~ L. 1989a. Ultrastructure comparCe de la spermiogenise et du spermatozoide de trois espZces de Pawvortex (Rhabdocoela, “Dalyellioida”, Graffillidae), turbellari& parasites intestinaux de mollusques. Zoologica Scripla 18: 161-174. NOURY-SRAIRI N., JUSTINE J.-L. & EUZET L. 1989b. Ultrastructure du ttgument de trois espices de Paravortex (Rhabdocoela, “Dalyellioida”, Graffillidae), turbellarits parasites intestinaux de mollusques. Annales des Sciences NaturelIes, Zoologie 10: 155-l 70. NOURY-SRAIRI N., JUSTINE J.-L. & EUZET L. 1989c. Implications phylogMtiques de l’ultrastructure de la spermatogentse, du spermatozoide et de I’ovogen&se du Turbellarik Urnstoma cyprinae (“Prolecithophora”, Urastomidae). Zooiogica Scripta 18: 161-174. NOURY-SRAIRI N., JUSTINE J.-L. & EUZET L. 1990. Ultrastructure du ttgument et des glandes sous-ipithbliales de Urastoma cyprinae (“Prolecithophora”), turbellari& parasite de mollusque. Annales des Sciences Nuturelles. Zoologie 11: 53-71. RIDLEY R. 1968. Electron microscopic studies on dicyemid Mesozoa. I. Vermiform stages. Journal of Parasitology54: 975-998. ROHDE K. 1987. Ultrastructural studies of epidermis, sense receptors and sperm of Craspedella sp. and Didymorchis sp. (Platyhelminthes, Rhabdocoela). Zoologicu Scripta 16: 289-295. ROHDE K. & WATSON N. 1988. Ultrastructure of epidermis and sperm of Syndisyrinx punicea (Hickman, 1956) (Rhabdocoela, Umagillidae). Ausrralian Journal qf Zoology 36: 131-139. ROHDE K., CANNON L.R.G. & WATSON N. 1988. Ultrastructure of the flame bulbs and protonephridial capillaries of Gieyszroria sp. (Rhabdocoela), Rhinolasius sp. (Rhabdocoela, Kalyptorhynchia), and Actinoductylelia blanchardi (Rhabdocoela, Temnocephalida). Journal of Submicroscopic Cyto1og.v and Pathology 20: 605-612.

Epidermis

of K. isopodicola

ROHDE K., WATSON N. & CANNON L. R. G. 1988. Comparative ultrastructural studies of the epidermis, sperm and nerve fibres of Cleistogamia longicirrus and Seritia stichopi (Rhabdocoela, Umagillidae). Zoologica Scripla 17: 337-345. ROHDE K. 1990. Phylogeny of Platyhelminthes, with special reference to parasitic groups. International Journal for Parasitology 20: 979-1007. ROHDE K., NOURY-SRAIRI N., WATSON N., JUSTINE J.-L. & EUZE~ L. 1990. Ultrastructure of the flame bulbs of Urastoma cyprinae (Platyhelminthes, ‘Prolecithophora’, Urastomidae). Acta Zoologica 71: 21 l-216. ROHDE K. 1991. The evolution of protonephridia of the Platyhelminthes. Hydrobiologia 227: 3 15-32 1. SCHOCKAERTE. R. & BEDINI C. 1977. Observations on the ultrastructure of the proboscis epithelia in Polycystis naegelii Kiilliker (Turbellaria Eukalyptorhynchia) and some associated structures. Acta Zoologica Fennica 154: 175-191. SMITH J. P. S. III, TYLER S., THOMAS M. B. & RIEGER R. M. 1982. The morphology of turbellarian rhabdites: phylogenetic implications. Transactions of the American Microscopical Society 101:209-228. TYLER S. 1976. Comparative ultrastructure of adhesive systems in the Turbellaria. Zoomorphologie 84: l-76. TYLER S. 1984. Turbellarian platyhelminths. In: Biology of the Integument, Vol. 1, Invertebrates (Edited by BEREITERHAHN J., MATOLTSYA. G. & RICHARDSK. S.), pp. 112-131. Springer, Berlin. WATSONN., ROHDE K. &WILLIAMS J. B. 1992. Ultrastructure of the protonephridial system of larval Kronborgia isopodicola (Platyhelminthes). Journal of Submicroscopic Cytology and Pathology 24: 43-49. WATSON N. A., WILLIAMS J. B. & ROHDE K. 1992. Ultrastructure and development of the eyes of larval Kronborgia isopodicola (Platyhelminthes, Fecampiidae). Acta Zoologica 73: 95-102. WILLIAMS J. B. 1975. Studies on the epidermis of Temnocephala I. Ultrastructure of the epidermis of Temnocephala novaezealandiae. Australian Journal of Zoology23: 321-331. WILLIAMS J. B. 1980a. Studies on the epidermis of Temnocephala V. Further observations on the ultrastructure of the epidermis of Temnocephala novaezealandiae, including notes on the glycocalyx. Australian Journal of Zoology 28: 43-57. WILLIAMS J. B. 1980b. Morphology of a species of Temnocephala (Platyhelminthes) ectocommensal on the isopod Phreatoicopsis terricola. Journal of Natural History 14: 183-199. WILLIAMS J. B. 1982. Studies on the epidermis of Temnocephala VI. Epidermal topography of T. novaezealandiae and other Australasian temnocephalids, with notes on microvillar and coated vesicle function and

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the evolution ofa cuticle. Australian Journal qf Zoology 30: 375-390. WILLIAMSJ. B. 1983. The genital system of Temnocephala I. Ultrastructural features of the differentiating spermatid of Temnocephala novaezealandiae, including notes on a possible correlation between cellular autophagy and mitochondrial function. Australian Journal of Zoology 31: 317-331. WILLIAMSJ. B. 1984. The genital system of Temnocephala II. Further observations on the spermatogenesis of Temnocephala novaezealandiae, with particular reference to the mitochondria. Australian Journal qf Zoology 32: 447-46 1. WILLIAMS J. B. 1986. Phylogenetic relationships of the Temnocephaloidea (Platyhelminthes). Hydrobiologia 132: 59-67. WILLIAMSJ. B. 1988a. Further observations on Kronborgia isopodicola, with notes on the systematics of the Fecampiidae (Turbellaria: Rhabdocoela). New Zealand Journal of Zoology 15:2 I IL22 1. WILLIAMSJ. B. 1988b. Ultrastructural studies on Kronborgia (Platyhelminthes: Neoophora): the spermatozoon. International Journalfor Parasitology 18:477-483. WILLIAMSJ. B. 1989. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae): the oocyte of K. isopodicola, with comments on oocyte microvilli and chromatoid bodies. International Journal for Parasitology 19: 207-216. WILLIAMSJ. B. 1990a. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae). The differentiated vitellocyte of K. isopodicola Blair and Williams. Australian Journal of Zoology 38: 79-88. WILLIAMSJ. B. 1990b. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae): epidermis and subepidermal tissues of the parasitic male K. isopodicola. International Journalfor Parasitology 20: 329-340. WILLIAMSJ. B. 1990~. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae): subepidermal glands of the female K. isopodicola. International Journal .for Parasitology 20: 803-8 13. WILLIAMSJ. B. 1990d. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae): observations on the genital system of the female K. isopodicola. International Joumalfor Parasitology 20: 957-963. WILLIAMSJ. B. 1991. Ultrastructural studies on Kronborgia (Platyhelminthes: Fecampiidae): observations on the encapsulated larva of K. isopodicola. New Zealand Journal of Zoology 18: 251-265. WILLIAMS J. B. 1992. Ultrastructure of the intestinal epithelium and prey microorganisms of Troglocaridicola mrazeki (Platyhelminthes: Temnocephaloidea), a scutariellid from Georgia, U.S.S.R. Journal of Submicroscopic Cytology 24: 47348 1. WILLIAMSJ. B. 1933. Nature, origin and fates ofmembranous lamellae in the lung of the neonate rat. Tissue and Cell 25: 481493.