Morphological features of the testis of freshwater blood flukes of the genus Sanguinicola Plehn, 1905, with consideration of the testicular patterns in the Aporocotylidae.

This is the first detailed study assessing the morphological features of the testis and testicular sperm of members of the freshwater blood flukes Sanguinicola sp. from Leuciscus idus (Cyprinidae) and for S. volgensis from Pelecus cultratus (Cyprinidae), and the marine blood fluke Aporocotyle simplex from Hippoglossoides platessoides (Pleuronectidae). The present study reports a unique feature in the distribution of germinal cellular components in freshwater Sanguinicola sp., showing the presence of the individual spermatocytes or their clusters in the testicular lobes, and the gathering of spermatid rosettes and spermatozoa within middle testicular lumen, which extends along the entire length of the testis. In contrast, each testis of marine A. simplex contains the usual mixed distribution of germ cells at various stages of development. The first TEM data on spermatozoon character of studied species has shown, unusual for digenean sperm structure, the absence of cortical microtubules in sperm principal region. Moreover, a variation in axoneme patterns is revealed in the studied aporocotylids, belonging to the different aporocotylid lineages a 9 + 0 axonemal type observed for freshwater teleost-infective species, Sanguinicola sp. and S. volgensis, and a 9 + '1' axonemal type revealed in spermatozoa of marine teleost-infective species A. simplex. The results discussed with the published data on the digenean sperm structure and the testicular patterns in the Aporocotylidae likely represent additional characteristics supporting the divergent evolutionary lineages of freshwater and marine aporocotylids. We anticipate future morphological studies of the sperm structure in aporocotylids of three lineages for an understanding of their phylogenetic relationships.

Are glial cells of the Digenea (Platyhelminthes) muscle cells?

Muscle cells of a digenean fish blood fluke, Aporocotyle simplex, aggregate along the periphery of the cerebral ganglia. Solitary myocytons and sarcoplasmic processes with muscle fibres give rise to long, narrow lamellate projections, which are visible along the periphery and within ganglia. These ultrastructural observations suggest a switching of glial functions to muscle cells and represent additional evidence of the phylogenetic lability of glial cells in bilaterians.

Heteromorphism of sperm axonemes in a parasitic flatworm, progenetic Diplocotyle olrikii Krabbe, 1874 (Cestoda, Spathebothriidea).

The spermatozoon ultrastructure of the progenetic cestode Diplocotyle olrikii (Spathebothriidea) has been examined using transmission electron microscopy and cytochemical staining with periodic acid-thiosemicarbazide-silver proteinate (PA-TSC-SP) for glycogen. The spermatozoon is a filiform cell, tapered at both extremities. Its moderately electron-dense cytoplasm possesses two parallel axonemes of unequal lengths. New for the Cestoda is a finding of three types of the mature spermatozoa with respect to different axonemal structure. The first type has both axonemes with standard 9 + '1' trepaxonematan pattern. The second type is represented by a spermatozoon having one axoneme with 9 + '1' structure and the second one with 9 + 0 pattern. The third type includes the two axonemes with 9 + 0 pattern. Microtubule doublets of the 9 + 0 axonemes contain either inner dynein arms or no dynein arms. In addition to the two axonemes, all three types of the mature sperm cells contain parallel nucleus, parallel cortical microtubules, four electron-dense plaques/attachment zones, and glycogen. The anterior extremity of the gamete exhibits a centriole surrounded by a semiarc of up to five electron-dense tubular structures. The distal end of the first type spermatozoa exhibits two morphological variants, represented either by (i) nucleus or (ii) remnants of the disorganized axoneme. Distal extremity of the spermatozoa of the second and third types contains doublets and singlets of disorganized axoneme. The ultrastructural characters of the spermatozoon of progenetic D. olrikii support the basal position of the Spathebothriidea within the Eucestoda.

Interrelationships of vitelline and muscle cells within the vitelline follicles of the blood fluke Aporocotyle simplex (Digenea, Aporocotylidae) and morphological evidence for the modification of vitelline material for eggshell formation.

This study of the fish blood fluke Aporocotyle simplex represents the first detailed transmission electron microscopical (TEM) investigation of the vitellarium of an aporocotylid digenean blood fluke. It revealed some unusual characteristics in the cytoarchitecture of the vitelline follicles and demonstrated modifications of the vitelline granules for eggshell formation. The vitelline follicles consist of vitellocytes at different developmental stages surrounded by sarcoplasmic processes of myocytes which occur throughout each follicle. Sites of intimate contact occur between the vitellocytes and the myocytes. Individual vitelline globules (0.1-0.2 µm in diameter) accumulate in quite small clusters of 10-20 and have a dense, heterogeneous matrix possessing central and peripheral regions with a greater density. Modifications of the vitelline globules take place within the clusters and are first apparent when the vitellocytes reach the lumen of the vitelline duct and vitelline reservoir. Globules within the clusters become confluent, and, when the vitellocytes reach the lumen of the oviduct and proximal ootype, these consolidated clusters contain a shapeless, loosely packed, dense material which is released from the vitellocytes by exocytosis. This investigation has provided morphological evidence for shell formation from modified vitelline globules in the form of a discontinuous, thin layer (~ 0.07 µm in thickness) of electron-dense shell material around the fertilized ovum and associated vitellocytes in the proximal ootype. The eggshell of intra-uterine eggs acquires an additional thin, heterogeneous outer layer, increasing its thickness to ~ 0.1 µm. The cytoarchitecture of the vitellarium, modifications of the vitelline globules within the clusters and the structure of the eggshell of A. simplex may prove to be of value in studies examining relationships between the three distinct lineages of aporocotylid digeneans.

Spermiogenesis produces the spermatozoa with 9 + '1' and 9 + 0 axonemal pattern in progenetic cestode Diplocotyle olrikii Krabbe, 1874 (Spathebothriidea: Acrobothriidae).

Spermiogenesis in the progenetic spathebothriidean cestode Diplocotyle olrikii has been examined using transmission electron microscopy (TEM) for the first time. Along with the typical features of spermatozoon cytodifferentiation (e.g., the electron-dense material in the apical region of the differentiation zone in the early stage of spermiogenesis, the intercentriolar body which is composed of three electron-dense plates and two electron-lucent zones, the orthogonal development of the two flagella, a flagellar rotation, proximo-distal fusion, the presence of two pairs of electron-dense attachment zones), new for the Eucestoda is detection of the formation of two types of free flagella during spermiogenesis in progenetic D. olrikii, exhibiting either standard 9 + '1' trepaxonematan pattern, or atypical 9 + 0 structure. Various combinations of these two types of flagella resulted in the production of three types of male gametes during spermiogenesis in this spathebothriidean cestode. The first type is represented with the two axonemes of the 9 + '1' structure; the second type exhibits two different axonemes, i.e., one with 9 + '1' and the other of 9 + 0 pattern; and the third type has two axonemes with atypical 9 + 0 structure. The occurrence of three sperm types in progenetic D. olrikii is associated with typical spermiogenesis and has never been described previously in the Platyhelminthes. We suppose that heteromorphism of male gametes in D. olrikii might be linked to progenesis, i.e., the programmed sexual maturation detected during the larval/developmental stage of an organism.

Unique ultrastructural characteristics of the tegument of the digenean blood fluke Aporocotyle simplex Odhner, 1900 (Digenea: Aporocotylidae), a parasite of flatfishes.

This paper includes the first transmission electron microscopical (TEM) study of the tegument of a member of the basal digenean family Aporocotylidae. Scanning electron microscopical investigations of the fish blood fluke Aporocotyle simplex show that each boss on the lateral body surface bears 12-15 simple, uniform spines which extend from 0.5-2.7 µm above the surface of the boss. TEM observations revealed that these spines reach deep beneath the distal cytoplasm of the tegument for much of their length (9-12 µm) and are surrounded by a complex of diagonal muscles in each boss. This is the first record of any digenean with so-called 'sunken' spines. The results suggest that aporocotylid spines arise from within the sarcoplasm of the boss diagonal muscles. The sunken cell bodies (perikarya) of the tegument are connected to the distal cytoplasm via ducts (specialised processes lined by microtubules); this in contrast to other digeneans studied, where they are connected via non-specialised cytoplasmic processes. Within the distal cytoplasm, the tegumental ducts of A. simplex are surrounded by invaginations of the basal membrane and release their cytoplasmic inclusions into the distal cytoplasm. These apparently unique morphological features of the tegument, especially the deep origin of the spines, may represent useful characteristics for understanding aporocotylid relationships, especially in view of the known variation in the spine patterns of aporocotylids.

First ultrastructural and cytochemical data on the spermatozoon and its differentiation in progenetic and adult Archigetes sieboldi Leuckart, 1878 (Cestoda, Caryophyllidea, Caryophyllaeidae).

Spermiogenesis in progenetic and adult stages of Archigetes sieboldi Leuckart, 1878, a tapeworm parasitic in oligochaetes and fish respectively, has been examined using transmission electron microscopy and cytochemical staining for glycogen. General pattern of spermiogenesis is essentially like that of other caryophyllideans, i.e., apical dense material in the zone of differentiation in the early stages of spermiogenesis, rotation of free flagellum and a flagellar bud, and proximo-distal fusion. Interestingly, rotation of a free flagellum and flagellar bud to the median cytoplasmic process (MCP) has been observed unconventionally at > 90° only in progenetic stages. Typical striated roots associated with the centrioles occur rarely in A. sieboldi, and only in form of faint structures in advanced stages of spermiogenesis. In contrast to most caryophyllideans studied to date, penetration of the nucleus into the spermatid body has started before the fusion of the free flagellum with the MCP. This feature has been reported rarely but exclusively in the family Caryophyllaeidae. The unipartite mature spermatozoon of A. sieboldi is composed of one axoneme of the 9 + '1' trepaxonematan pattern with its centriole, parallel nucleus, and parallel cortical microtubules which are situated in a moderately electron-dense cytoplasm with glycogen particles. An unusual arrangement of cortical microtubules in the two parallel rows in region I of the spermatozoon is described here for the first time in the Caryophyllidea. Ultrastructural data on spermiogenesis and the spermatozoon in A. sieboldi from tubuficids and carp are compared and discussed with those in other caryophyllideans and/or Neodermata.

The first ultrastructural observations of the egg-forming complex of Calicotyle affinis, an endoparasitic monocotylid monogenean which lacks a uterus.

This ultrastructural study of the female reproductive system of Calicotyle affinis, a monogenean without a uterus, demonstrates, for the first time in a monogenean, the presence of accessory cells located in the peripheral region of the ovary and a single large cell whose surface is penetrated by deep invaginations which restrict the passage of oocytes through the lumen of the distal extremity of the ovary. The cytoarchitecture of the epithelial lining of the two vaginae is a syncytium formed by an anucleate epithelial lining with sunken epithelial perykaria. The wall of the seminal receptacle is an enlarged prolongation of the vaginal epithelial lining. Fertilization occurs in the fertilization chamber and fertilized oocytes retain cortical granules within their cytoplasm, a characteristic known to occur in free-living flatworms. Our study also highlights concentrations of two distinct groups of Mehlis' gland cell ducts on either side of the proximal end of the ootype distinguished by ultrastructural characteristics of their secretory granules. The epithelial wall of the ootype is formed by a single layer of regular, columnar, glandular epithelial cells; these cells are closely adjacent, conjoined towards their apical region by septate junctions and produce rounded, electron-dense granules which are discharged into the ootype lumen via a merocrine or holocrine mechanism. Released granules concentrate around the eggshell and form an additional fibrous coat. The morphological diversity observed in the female reproductive system of the Monogenea is commented on it relation to clarifying patterns in monogenean evolution and for understanding the phylogeny of the Neodermata.

Ultrastructural characteristics of the male ducts and terminal genitalia of an endoparasitic monogenean, Calicotyle affinis Scott, 1911 (Monopisthocotylea: Monocotylidae), with the first detailed description of a copulatory stylet in a monogenean.

This investigation of Calicotyle affinis is the first ultrastructural description of the male reproductive system of a monocotylid monogenean. It has revealed numerous characteristics which are either new or clarify those determined from previous light microscopical observations on the morphology of Calicotyle spp. These include numerous individual testes, each surrounded by its own basal lamina; an ejaculatory bulb delimited by a basal lamina surrounded by two independent internal and external bands of muscles; a bipartite, but not fully divided, muscular chamber (an internal seminal vesicle) in the proximal region of the ejaculatory bulb; well-developed unicellular prostatic glands localised outside the ejaculatory bulb, the cytoplasmic extensions of which pass into the ejaculatory bulb, to form the ducts which open into the lumen of ejaculatory duct; and a male copulatory organ in the form of a coiled penis tube containing throughout its lumen a long, hollow stylet. The penis tube appears to be a modified prolongation of the ejaculatory duct and is supported by a basal lamina and muscle layers. The stylet wall consists of a homogenous, dense material with a narrow, denser inner lining; it is funnel-shaped proximally but narrows imperceptibly towards the distal end of the penis tube, and appears to be a derivative of the basal laminae of the ejaculatory bulb and distal ejaculatory duct. A comparative analysis of the nature of the stylet in relation to those found in free-living platyhelminths forms part of a search for characters useful for understanding monocotylid origins and relationships.

The unique uterine structure of the basal monogenean Chimaericola leptogaster (Monogenea: Polyopisthocotylea), an ectoparasite of the relictual holocephalan fish Chimaera monstrosa.

Ultrastructural studies of the monogenean uterus are few in number and no non-polystomatid polyopisthocotyleans have been investigated. The uterus of Chimaericola leptogaster, a basal polyopisthocotylean monogenean, has several unusual features, including six reflexed loops comprising four ascending and three descending, longitudinally oriented, linear sections. At the ultrastructural level, three readily distinguishable uterine regions and other distinctive characteristics are apparent. One novel feature occurs in the proximal uterus, where the lining forms a so-called 'single-layered multi-rowed cellular epithelium', which includes two types of cells, tall (ca. 14-19 µm in height) and short (ca. 6-9 µm in height) cells, both lying on the basement membrane. Although known from other bilaterian groups, this is the first record of this type of epithelium in the Neodermata. The lining of the middle uterine region comprises a single regular layer of columnar glandular epithelial cells, which produce numerous rounded, electron-dense granules associated with Golgi complexes. The presence of the uterine glands in the middle region of the uterus is an unusual feature for a monogenean, having previously been described only for basal orders of the Cestoda, i.e. the Gyrocotylidea, Caryophyllidea and Spathebothriidea. Seen in cross-section, the epithelium of the distal uterus contains three areas of tall single-layered columnar epithelium (ca. 30 µm deep) interspersed by three areas of flattened epithelium (ca. 0.2-0.9 µm deep). Such a pattern is quite different from those reported for other monogeneans and, indeed, other neodermatan groups. The investigation has shown that the outer layer of the fully formed eggshell is assembled from epithelial secretions in the middle uterine lumen, but is modified in terms of its shape in the distal uterus. Possible phylogenetic implications arising from the unusual features described are discussed in relation to other neodermatan groups and recent molecular phylogenies of the Bilateria.

Surface ultrastructural characteristics of Dictyocotyle coeliaca Nybelin, 1941 (Monopisthocotylea: Monocotylidae), an endoparasitic monogenean of rays.

Scanning and transmission electron microscopical observations were made of the surface topography of Dictyocotyle coeliaca (Nybelin, 1941) (Monopisthocotylea: Monocotylidae), a unique endoparasitic monogenean from the body cavity of the ray Amblyraja radiata (Elasmobranchii: Rajidae). Scanning investigation show the presence on the ventral side of the anterior body of smooth areas with pit-like depressions and shallow ridges, whereas the tegument of the middle and posterior regions of the body is extensively folded, and on the dorsal surface the tegument is smooth but interrupted by deep depressions. Transmission observation revealed the presence of invaginations of varying irregularity and depth, which form the various depressions of the tegumental surface. Non-ciliated, dome-shaped papillae occur singly or in groups and are common around the mouth and both the genital and vaginal pores, but exhibit no particular orientation. The haptor is much reduced, compared with that of ectoparasitic monocotylids, and is covered ventrally with an irregular array of 40-70 shallow loculi. Internal differences occur in the thickness of the syncytial tegumentary layer of the haptoral loculi and septa and also in the number of cytoplasmic inclusions. The locular surface has distinct ultrastructural characteristics, the most obvious of which are a honeycomb arrangement of small pockets measuring 0.6 × 0.75 to 1.0 × 1.7 µm in diameter, a terminal web beneath the surface plasma membrane and large outgrowths on the surface of the loculi filled with lysosome-like bodies. The surface specializations of the body and haptoral tegument of D. coeliaca are discussed in relation to their being adaptations to an endoparasitic environment.

Ultrastructural observations of the attachment organs of the monogenean Rajonchocotyle emarginata (Olsson, 1876) (Polyopisthocotylea: Hexabothriidae), a gill parasite of rays.

The present study uses scanning and transmission electron microscopical observations to examine the haptoral features of the hexabothriid polyopisthocotylean monogenean Rajonchocotyle emarginata from the gills of the elasmobranch Amblyraja radiata. The haptor possesses six equal, pedunculate suckers, each armed with a large, curved, hooked sclerite, and an appendix bearing two small, terminal suckers and a pair of minute hamuli. The outer side of the distal region of the hooked sclerite forms a large bulge along the antero-lateral side of each sucker and the proximal region forms a small bulge on the opposite postero-lateral side; part of the large bulge, which covers the small sclerite hook, surmounts the sucker rim. The sucker sclerite is situated outside the main sucker muscle complex of the sucker wall and is surrounded by its own muscular envelope; close to the sclerite itself, long, tightly packed lamella-like processes of sarcoplasm are present. A muscular sphincter surrounds the distal region of the sclerite shaft. The sclerite consists of three heterogeneous layers, but its hook region is distinguished from the shaft by the presence of an additional homogeneous layer. Hamuli within the appendix consist of three distinct layers loosely surrounded by sarcoplasmic lamellae. The luminal surface of the appendicular suckers is covered with unusual, long, thin, interconnected surface protrusions. In all other regions of the haptor, the tegumental surface is smooth, with numerous shallow pores penetrating the "terminal web" of the syncytial cytoplasm. Unicellular gland cells are localized close to the base of the appendix, with their ducts, containing non-homogenous secretory bodies, opening onto the anterior haptoral surface. Two types of sense receptors are visible on the haptoral surface. The ultrastructural features of the haptor are discussed in relation to our understanding of their function and the evolutionary relationships of the basal polyopisthocotylean monogenean groups Hexabothriidae and Chimaericolidae.

Ultrastructural characteristics of the caeca of basal polyopisthocotylean monogeneans of the families Chimaericolidae and Hexabothriidae parasitic on cartilaginous fishes.

Ultrastructural differences are shown between the caecal organization in three blood-feeding polyopisthocotylean monogeneans, i.e., the chimaericolid Chimaericola leptogaster from the holocephalan Chimaera monstrosa and two hexabothriids, Callorhynchocotyle callorhynchi from the holocephalan Callorhynchus capensis and Rajonchocotyle emarginata from the elasmobranch Amblyraja radiata. In C. leptogaster, digestive cells and connecting syncytium, joined close to the luminal surface by septate junctions, are arranged alternately along the caecal epithelial wall; the nuclear regions of both cell types are sunk below the general level of the caecal epithelium; a concave depression on the apical margin of the digestive cells bears lamellae; and this depression is covered by a lamellate bubble formed by thin projections emanating from the connecting syncytium. The luminal surface of the connecting syncytium is covered with outgrowths terminating in the form of long, narrow processes. In R. emarginata and C. callorhynchi, the predominant digestive cells are at different stages of development and occur in groups, developing digestive cells bulge into the caecal lumen from the connecting syncytium with contact sites present close to the luminal surface, and the luminal surface structures of both the connecting syncytium and the digestive cells are short lamellae. In these two hexabothriids, a holocrine (or apocrine) process for the elimination of digestive product is assumed via the detachment of fully differentiated, bulging digestive cells. Free, apparently sloughed digestive cells and residual bodies are present within the caecal lumen, and replacement digestive cells are numerous in the connecting syncytium. In the chimaericolid, free bubbles containing residual bodies and portions of digestive cells filled with degenerating digestive vesicles occur in the caecal lumen along with large amounts of male and female reproductive material. The usefulness of characteristics of the caecal ultrastructure as taxonomic traits at the family level is discussed.

The surface topography of Callorhynchocotyle callorhynchi (Manter, 1955) (Monogenea: Hexabothriidae), a parasite of the holocephalan fish Callorhinchus capensis.

A scanning electron microscopical study, incorporating some transmission electron microscopical observations, was undertaken on the surface topography of the gill parasite Callorhynchocotyle callorhynchi (Manter, 1955) (Monogenea: Hexabothriidae) from the Cape elephant fish Callorhinchus capensis (Holocephali) off the western coast of South Africa. The study revealed the presence of several new characteristics for this species. These include the presence of regularly distributed, knob-shaped projections on the surface of the haptor, haptoral appendix and sucker peduncles measuring 0.2 µm and in concentrations of approximately 100 per 10 µm(2) and the existence of a ridge which bisects each sucker lumen, forming two different loculi. We also report, for the first time for any monogenean, the presence of 'true spines'; these occur on the luminal surface of the haptoral suckers and have all of the characteristics of the tegumental spines of digeneans, i.e. they are situated within the distal syncytial tegumental cytoplasm, rest on the basal plasma membrane, have a uniform structure and are covered apically by the tegumental surface plasma membrane. These spines are simple, straight and single-pointed. Under the scanning electron microscopy (SEM), within an area of 20 µm in diameter, 23 such spines were counted, but their concentration and arrangement varies in different regions of the sucker. At their base, they measure about 1.5 µm in width and reach approximately 2 µm in height above the general level of the tegument, but transmission electron microscopy (TEM) measurements of an entire spine indicate that they may reach 3 µm in total length. The presence of spines, possessing similar morphological characteristics in both basal polyopisthocotylean monogeneans and digeneans, represents another characteristic which may prove useful in understanding the evolutionary relationships within the Neodermata.

Ultrastructure of the anterior organ and posterior funnel-shaped canal of Gyrocotyle urna Wagener, 1852 (Cestoda: Gyrocotylidea).

Using scanning and transmission electron microscopy, ultrastructure of the anterior organ and posterior funnel-shaped canal of Gyrocotyle urna Wagener, 1852 (Cestoda: Gyrocotylidea) from ratfish, Chimaera monstrosa (Holocephali), was studied for the first time. The proper anterior organ is localised at a short distance (about 170 µm) from an apical pore surrounded by a receptor field, whereas its distal end is marked by a muscular sphincter. The tegumental surface of this organ is covered with short filitriches of irregular length; large area of muscle layers traverse beneath the tegumental layer. The funnel-shaped canal of G. urna (2.5-3.0 mm long) is a specialised, muscular part of the posterior attachment organ; it opens on the rounded elevation on the dorsal body surface. The tegumental layer bears conical sclerite-like structures (up to 1.5 µm long). It produces electron-dense bodies that are transported into a canal lumen and surrounded thick muscle area mixed with numerous nerve fibres. The present ultrastructural study of G. urna indicates that gyrocotylideans share some ultrastructural characters of the anterior organ with spathebothriidean cestodes with a single anterior attachment sucker-like organ. In contrast, the unique posterior rosette attachment organ with funnel-shaped canal of the Gyrocotylidea resembles the haptor of polyopisthocotylean monogeneans in its position at the posterior end of the body and presumed origin. The above-mentioned features add more clarity to support the basal position of the Gyrocotylidea Poche, 1926 among cestodes. In addition, they also indicate a possible relationship of gyrocotylidean ancestors with monogeneans.

Clamp ultrastructure of the basal monogenean Chimaericola leptogaster (Leuckart, 1830) (Polyopisthocotylea: Chimaericolidae).

The ultrastructure of the haptoral clamps of the chimaericolid monogenean Chimaericola leptogaster, a basal polyopisthocotylean from the gills of a holocephalan fish, is described. These clamps are characterized by the presence of two muscle blocks interrupted mid-anteriorly and mid-posteriorly and different kinds of hard structures: a single median and paired lateral sclerites embedded in the clamp wall; six spine-like structures directed towards the clamp lumen; and electron dense surface structures along the internal surface of the anterior clamp lips and along the luminal surface of the tegument of the clamp lumen. The lateral sclerites are situated deep within muscular tissue and are closely bounded by radial myofibrils, possessing a uniform electron dense matrix within which are hollow areas of different sizes. The median sclerite occupies an area between the clamp wall myofibrils and the luminal epithelium, is surrounded by a basement lamina and is composed of a heterogeneous matrix comprising two different morphological layers related to variations in the type and concentration of fibrils. Four of the spine-like structures are extensions of the margins of the two spindle-like muscle blocks in the clamps, i.e. the two anterior and two posterior structures, and the two others are situated at the lateral constrictions of the left and right muscle blocks. The electron dense surface structures are derivations of the clamp tegument or, to be more precise, its outer, densely fibrous region. These results are discussed in relation to the evidence that the haptoral clamps of C. leptogaster are apparently ancient origin.

Ultrastructural features of the spermatozoon and its differentiation in Brandesia turgida (Brandes, 1888) (Digenea, Microphalloidea, Pleurogenidae).

Spermatological characters of the digenean Brandesia turgida (Brandes, 1888), an intestinal parasite of the frog Pelophylax ridibundus (Pallas, 1771), have been investigated by means of transmission electron microscopy for the first time. The process of the spermatozoon formation begins with the appearance of the differentiation zone bordered by cortical microtubules and containing two centrioles associated with striated rootlets and with an intercentriolar body. The intercentriolar body is made up of seven distinct electron-dense plates, two less electron-dense, and four electron-lucent zones. The orthogonal development of the two flagella is followed by a flagellar rotation and their proximodistal fusion with the median cytoplasmic process. This process is accompanied by an extension of both the mitochondrion and nucleus into the median cytoplasmic process. The mature spermatozoon of B. turgida contains two parallel axonemes of unequal lengths with the 9 + "1" trepaxonematan pattern, mitochondrion, nucleus, parallel cortical microtubules, four electron-dense attachment zones, an external ornamentation of the plasma membrane, and electron-dense glycogen granules. The anterior extremity of the male gamete contains one complete centriole, a small component of the central element of the second centriole, and peripheral cortical microtubules (up to 45). The posterior extremity of the mature spematozoon exhibits tubular elements of the disorganized axoneme. The present study provides the first data on spermiogenesis within the family Pleurogenidae. Variations of the spermatozoa ultrastructural characters within Digenea, in particular, between different families of the superfamily Microphalloidea, are discussed.

Ultrastructural evidence for completion of the entire miracidial maturation in intrauterine eggs of the digenean Brandesia turgida (Brandes, 1888) (Plagiorchiida: Pleurogenidae).

Results of this TEM study provide ultrastructural evidence that miracidial morphogenesis is fully completed within the intrauterine eggs situated in the most posterior uterine regions of the pleurogenid trematode Brandesia turgida (Brandes, 1888). The ultrastructural characteristic of different larval organelles and cell types of these eggshell-enclosed, but fully formed, cilated miracidia is described. The body wall of the pyriform mature miracidium of B. turgida is composed of ciliated epidermis and underlying peripheral body musculature. Two miracidial flame cells of the protonephridial excretory system are localized in the central region of the ciliated larvae. Three types of miracidial glands were observed: a single apical gland, two lateral glands, and several small vesiculated glands; each gland type contains characteristic, but different types of secretory granules. The anterior end of each miracidium consists of an apical papilla on which are situated the exits of the three main larval glands: an exit of a single apical gland as well as the individual exits of two lateral glands. The exits of vesiculated glands, containing characteristic spherical membrane-bound and highly electron-dense granules, evidently different from the two other types of secretory granules of apical and lateral glands, were not identified. Germinative cells, grouped together in a sac-like germinative follicle, are situated in the medioposterior part of the larva, the germatophore. The germinative cells contain numerous electron-dense heterochromatin islands arranged in the form of a network or chain-like pattern and distributed mainly in the karyoplasm adjacent to the nuclear membrane. The thin layer of granular cytoplasm is rich in free ribosomes and contains a few small mitochondria. Both nuclear and cytoplasmic features if these cells indicate their great developmental potential for further growth and multiplication in postembryonic stages of the life cycle. In the mature eggs, the areas of focal cytoplasmic degradation were frequently observed and may be involved in the autolysis of some embryonic structures. Obtained results are compared with available literature data on the functional ultrastructure of the miracidia of other digeneans.

Ultrastructure and chemical composition of the proboscis hooks of Acanthocephalus lucii (Müller, 1776) (Acanthocephala: Palaeacanthocephala) using X-ray elemental analysis.

The ultrastructure and chemical composition of the proboscis hooks and surrounding tegument of Acanthocephalus lucii (Müller, 1776), a parasite of European perch, Perca fluviatilis Linnaeus, were examined using scanning (SEM) and transmission (TEM) electron microscopy and X-ray microanalysis (EDXA). The blade of middle hooks consists of three layers: an outer homogeneous layer, an inner heterogeneous layer and a central core. TEM observation revealed the presence of hollow tubes, which spaced the central core; fibrous inner hook layer surrounded by an electron-dense margin and the basal tegumental layer filled with electron-dense bodies and outer layer. We found for the first time that the so-called 'epidermal covering' surrounding of the exposed hook blade (outer hook layer) is a modified striped portion of the tegumental layer and there are no special contact sites between these two morphologically different structures, i.e. striped layer of the syncytial tegument and following proper outer hook layer, which is a homogeneous, moderately electron-dense layer of -0.3 µm in thickness. The hook root is embedded into subtegumental fibrous layer. X-ray microanalysis of both the surface and internal parts of A. lucii hooks demonstrated the presence of calcium, magnesium, phosphorus and sulphur. The highest concentration of sulphur was recorded at the tip of hooks, whereas the middle part of the hooks was most rich in calcium, phosphorus and magnesium. The proximal part of the hooks contained lower concentrations of sulphur, calcium and phosphorus. In the proboscis tegument, only two elements, calcium and silicon, were found. The differences observed in the chemical composition of the hook 'epidermal covering' and the proboscis tegument support our ultrastructural findings that the hook tegumental covering is a modified structure compared with that of the general proboscis tegument.

Corrigendum: Trophic diversification and parasitic invasion as ecological niche modulators for gut microbiota of whitefish.

[This corrects the article DOI: 10.3389/fmicb.2023.1090899.].