Ultrastructure of Sperm in <Emphasis Type="Italic">Mercenaria stimpsoni</Emphasis> and <Emphasis Type="Italic">Mactra chinensis</Emphasis> (Mollusca: Bivalvia) from the Sea of Japan

The ultrastructure of the sperm of the common bivalve species Mercenaria stimpsoni and Mactra chinensis from Peter the Great Bay is described. The sperm structure is typical for animals with external insemination. The sperm consists of a head, middle part, and flagellum. The sperm head of M. stimpsoni has a curved crescent form and includes the nucleus and acrosome; the head length is 9.8 μm. The acrosome is subdivided to the acrosome granule and the periacrosomal material. There are 4 mitochondria of about 0.8 μm in size in the middle part of the spermatozoon. The mitochondria surround the centriolar apparatus, which consists of proximal and distal centrioles located at a right angle. The axoneme originates from the distal centriole. The sperm of M. chinensis is barrel-shaped, with a head length of 3.2 μm. The acrosome is relatively larger, and its height is 1–1.2 μm. There are also 4 mitochondria 0.6–0.8 μm in the middle part of the spermatozoon. The sperm structure of the described species is typical of the families to which the mollusks belong, with insignificant variations.     

Spermatozoa and spermatogenesis in the northern quahaug <Emphasis Type="Italic">Mercenaria mercenaria</Emphasis> (Mollusca,Bivalvia)

We studied the ultrastructure of spermatogenesis and spermatozoa in the northern quahaug, the clam Mercenaria mercenaria. Spermatogenetic cells gradually elongate. Mitochondria gradually fuse and increase in size and electron density. During spermatid differentiation, proacrosomal vesicles migrate towards the presumptive anterior pole of the nucleus and eventually form the acrosome. The spermatozoon of M. mercenaria is of a primitive type. It is composed of head, mid-piece, and tail. The acrosome shows a subacrosomal space with a short conical contour. The slightly curved nucleus of the spermatozoon contains fine-grained dense chromatin. The middle piece consists of a centriolar complex which is surrounded by four mitochondria. The flagellum has a standard “9 + 2” microtubular structure. The ultrastructure of spermatozoa and spermatogenesis of M. mercenaria shares a number of features with other species of the family Veneridae. M. mercenaria may be a suitable model species for further investigations into the mechanisms of spermatogenesis in the Bivalvia.     

The spermiogenesis and the early spermatozoa of <Emphasis Type="Italic">Armorloricus elegans</Emphasis> (Loricifera,Nanaloricidae)

The spermiogenesis consisting of five spermatid stages and the early spermatozoon has been investigated in Armorloricus elegans (Loricifera) with the use of transmission electron microscopy. The male reproductive system consists of three parts; testes, vasa deferentia and seminal vesicles. Caudally, the two seminal vesicles merge together in a ciliated duct and the excretory/gonadal—and digestive systems continue through the recto-urogenital canal, which opens via the lateral gonopores and the temporarily closed anal system. Spermiogenesis mainly occurs in the testes, whereas further maturation of the late spermatids and early spermatozoa occurs in the vasa deferentia and seminal vesicles. A maturation gradient (from spermatocytes to spermatozoa) is found from the posterior peripheral part of the testes to the anterior periphery and then centrally. During spermiogenesis the round nucleus becomes more osmiophilic and condensation of chromatin occurs. Later the nucleus elongates until it becomes rod-shaped in the early spermatozoa. In the second spermatid stage, a large vesicle is formed by saccules developed from the Golgi complex. This vesicle develops further and consists of three different osmiophilic parts with some crystal-like structures inside and is on the outside almost entirely surrounded by thick striated filaments. In the mid-piece the flagellum has a typical 9 × 2 + 2 axoneme and the two mitochondria are fused into a single sheet surrounding the flagellum. In the early spermatozoon stage an acrosomal-like cap structure with an acrosome filament appears proximal to the protruded rod-shaped nucleus. This cap is not formed by the Golgi complex and therefore might not be a true acrosome. Comparing the early spermatozoa of A. elegans with other cycloneuralians has shown some similarities with especially Kinorhyncha and Priapulida. These similarities are thought to be plesiomorphic.     

An ultrastructural comparative study of the sperm of<Emphasis Type="Italic"> Hyla pseudopseudis</Emphasis>,<Emphasis Type="Italic"> Scinax rostratus</Emphasis>, and<Emphasis Type="Italic"> S. squalirostris</Emphasis> (Amphibia: Anura: Hylidae)

We make detailed comparisons of the ultrastructure of the spermatozoon among three species of the family Hylidae, Hyla pseudopseudis, Scinax rostratus, and S. squalirostris. The acrosome complex consists of two conical structures covering the nuclear rostrum, the acrosome vesicle, and the subacrosomal cone. The nucleus has a moderately condensed chromatin with a conical shape in longitudinal sections and a circular shape in cross-sections. In H. pseudopseudis, mitochondria are numerous and circular, and in S. rostratus and S. squalirostris there are fewer mitochondria that are more elongate in longitudinal and transverse sections. In H. pseudopseudis, the mitochondrial collar starts adjacent to the distal centriole, occupying the whole midpiece, whereas in both Scinax species the mitochondrial collar starts only at the posterior one-third of the midpiece. In both Scinax species, the presence of juxta-axonemal fiber, axial sheath, and axial fiber in the tail are seemingly plesiomorphic characters, widespread among bufonoid frogs. In H. pseudopseudis, however, the absence of axial fiber and axial sheath seems to be derived from the typical bufonoid condition. The differences between Hyla and Scinax sperm endorse the separation of the two genera and suggest that sperm ultrastructure can be a useful tool to investigate relationships at the intrafamily level.     

Fine structure of the spermatozoon of <Emphasis Type="Italic">Diopatra neapolitana</Emphasis> (Polychaeta,Onuphidae)

The identification of Diopatra species lacks of clear diagnostic features of taxonomic importance and the knowledge of their reproductive characters is scant. The spermatozoa of Diopatra neapolitana were ultrastructurally investigated by electron microscopy in order to correlate the mode of reproduction with sperm cells morphology. The mature male gamete has a depressed subspherical nucleus, a cone-like acrosome, and a long flagellum. The acrosome is conical in shape and radially symmetrical, with a base diameter twice the height. Within the acrosome vesicle, the basal region includes a very electron-dense thickened ring composed of paracrystalline substances. The subacrosomal space is filled with a poorly electron-dense material, with straight filaments axially arranged to form a perforatorium. The nucleus contains the complete axial canal, holding the hind perforatorium region. The middle piece consists of five mitochondria with well-distinct membranes and tubulo-vesicular cristae. Two centrioles are located perpendicularly to each other. The proximal one lies in the central fossa and the distal one, slightly eccentric to the sperm axis, anchors to the plasma membrane by nine satellite rays of the pericentriolar complex. The axoneme has a 9+2 arrangement of microtubules. In general, the spermatozoon of D. neapolitana conforms exteriorly to the typical ect-aquasperm; the acrosome complex ultrastructure, however, shows noticeable modifications from the basic form. This finding agrees with the previously observed reproductive pattern (broadcast spawning—free-swimming larvae) of D. neapolitana belonging to Santa Gilla population, and may be helpful to solve the taxonomic problems of the D. neapolitana complex as well.     

Sperm ultrastructure of<Emphasis Type="Italic"> Mantophasma zephyra</Emphasis> (Insecta,Mantophasmatodea)

The ultrastructure of Mantophasma zephyra spermatozoa is described. Sperm cells have a trilayered acrosome with conspicuous extra-acrosomal material which expands along the nucleus. The nucleus is crossed anteriorly by a canal and its posterior end is embedded in the centriole adjunct material. A centriole with microtubular triplets is present. The flagellum has a 9+9+2 axonemal pattern, two partially crystallised mitochondrial derivatives, two membranous sacs and three connecting bands. The accessory microtubules are filled with dense material and have 16 protofilaments in their tubular wall. The intertubular material is not very expanded. In the seminal vesicles spermatozoa are stuck together to form spermatodesms, and their heads are also joined by adherens junctions. A cladistic analysis based on sperm features indicates a close relationship of Mantophasmatodea with Mantodea.     

Ultrastructural study of spermatogenesis in<Emphasis Type="Italic"> Phoronopsis harmeri</Emphasis> (Lophophorata,Phoronida)

The process of sperm development in Phoronopsis harmeri was studied by electron microscopy. Developing spermatogenical cells are aggregated around the capillaries of the haemal plexus. The spermatogonia, which are situated around the capillary walls of the caeca, are remarkable for the presence of germ-line vesicles and contain their centrioles near the cell membrane. The spermatocytes and spermatids are flagellated cells arranged in clusters. During spermiogenesis the basal body/flagellum complex migrates to the apical pole of the spermatid. The acrosome-like structure arises from material produced by the Golgi complex. It lacks a surrounding membrane and has a fibrillar content. The nucleus elongates and the condensation of chromatin is caused by an activation of 'initiation centres'. The late spermatid and the spermatozoon appear as two-armed 'V'-shaped cells in which one arm contains the nucleus and posteriorly located mitochondria, and the other one is the axoneme. Spermatogenesis of P. harmeri is an interesting example of gamete differentiation where advanced sperm structure is combined with a plesiomorphic pattern of sperm development characterized as 'flagellate spermatogenesis'. Communicated by H.-D. Franke     

Ultrastructural observations on the spermatozoa of a species of <Emphasis Type="Italic">Lepidodasys</Emphasis> (Gastrotricha,Macrodasyida)

The ultrastructure of the spermatozoon of a species in the marine gastrotrich genus Lepidodasys is described. The filiform cell is composed of a cork-screw acrosome, a long single mitochondrion surrounded by a helical nucleus, and a flagellum with a 9 × 2 + 2 axonemal arrangement. The structure of the sperm of this species from Denmark appears closely similar to those of the other two species of Lepidodasys studied so far from Italy and Florida (US). Peculiar features (cylindrical nucleus, absence of a periaxonemal sheath) place this genus far from the others in the family Lepidodasyidae. The absence of synapomorphies between Lepidodasys and other genera of Lepidodasyidae suggests that the family is polyphyletic. The sperm ultrastructure fully fits the species of Lepidodasys into the marine order Macrodasyida, with the sperm ground plan of which its sperm shares a number of details.     

Fine structure of acrosome biogenesis and of mature sperm in the bivalve molluscs<Emphasis Type="Italic"> Glycymeris</Emphasis> sp. (Pteriomorphia) and<Emphasis Type="Italic"> Eurhomalea rufa</Emphasis> (Heterodonta)

Proacrosomal vesicles form during the pachytene stage, being synthetized by the Golgi complex in Glycymeris sp., and by both the Golgi and the rough endoplasmic reticulum in Eurhomalea rufa. During early spermiogenesis, a single acrosomal vesicle forms and its apex becomes linked to the plasma membrane while it migrates. In Glycymeris sp., the acrosomal vesicle then turns cap-shaped (1.8 μm) and acquires a complex substructure. In E. rufa, proacrosomal vesicles differentiate their contents while still at the premeiotic stage; as the acrosomal vesicle matures and its contents further differentiate, it elongates and becomes longer than the nucleus (3.2 μm), while the subacrosomal space develops a perforatorium. Before condensation, chromatin turns fibrillar in Glycymeris sp., whereas it acquires a cordonal pattern in E. rufa. Accordingly, the sperm nucleus of Glycymeris sp. is conical and elongated (8.3 μm), and that of E. rufa is short and ovoid (1.1 μm). In the midpiece (Glycymeris sp.: 1.1 μm; E. rufa: 0.8 μm), both species have four mitochondria encircling two linked orthogonal (Glycymeris sp.) or orthogonal and tilted (30–40°; E. rufa) centrioles. In comparison with other Arcoida species, sperm of Glycymeris sp. appear distinct due to the presence of an elongated nucleus, a highly differentiated acrosome, and four instead of five mitochondria. The same occurs with E. rufa regarding other Veneracea species, with the acrosome of the mature sperm strongly resembling that of the recent Mytilinae. Electronic Publication     

Sperm structure of Mecoptera and Siphonaptera (Insecta) and the phylogenetic position of<Emphasis Type="Italic"> Boreus hyemalis</Emphasis>

Sperm ultrastructure has been studied in three species of the taxa Mecoptera and Siphonaptera. The spermatozoon of the scorpion fly Panorpa germanica shows an apical bilayered acrosome, a helicoidal nucleus, a centriolar region and a 9+2 flagellar axoneme helicoidally arranged around a long mitochondrial derivative. A second mitochondrial derivative is very short and present only in the centriolar region. A single accessory body is present and it is clearly formed as a prolongation of the centriole adjunct material. Two lateral lamellae run parallel to the nucleus. The snow fly Boreus hyemalis has a conventional sperm structure and shows a bilayered acrosome, a long nucleus, a centriolar region, two mitochondrial derivatives and two accessory bodies. The axoneme is of the 9+2 type and is flattened at the tail tip. Both P. germanica and B. hyemalis have two longitudinal extra-axonemal rods and have a glycocalyx consisting of longitudinal parallel ridges or filaments. The spermatozoon of the flea Ctenocephalides canis has a long apical bilayered acrosome, a nucleus, a centriolar region, a 9+2 axoneme wound around two unequally sized mitochondrial derivatives, and two triangular accessory bodies. In the posterior tail end the flagellar axoneme disorganises and a few microtubular doublets run helicoidally around the remnant mitochondrial derivative. The glycocalyx consists of fine transverse striations. In all three species, the posterior tail tip is characterised by a dense matrix embedding the disorganised axoneme. From this comparative analysis of the sperm structure it is concluded that Mecoptera, as traditionally defined, is monophyletic and that B. hyemalis is a member of Mecoptera rather than of Siphonaptera.     

Spermiogenesis and chromatin condensation in the common tree shrew, <Emphasis Type="Italic">Tupaia glis</Emphasis>

We have investigated the cellular characteristics, especially chromatin condensation and the basic nuclear protein profile, during spermiogenesis in the common tree shrew, Tupaia glis. Spermatids could be classified into Golgi phase, cap phase, acrosome phase, and maturation phase. During the Golgi phase, chromatin was composed of 10-nm and 30-nm fibers with few 50-nm to 60-nm knobby fibers. The latter were then transformed into 70-nm knobby fibers during the cap phase. In the acrosome phase, all fibers were packed into the highest-order knobby fibers, each about 80–100 nm in width. These chromatin fibers became tightly packed in the maturation phase. In a mature spermatozoon, the discoid-shaped head was occupied by the acrosome and completely condensed chromatin. H3, the core histone, was detected by immunostaining in all nuclei of germ cell stages, except in spermatid steps 15–16 and spermatozoa. Protamine, the basic nuclear protein causing the tight packing of sperm chromatin, was detected by immunofluorescence in the nuclei of spermatids at steps 12–16 and spermatozoa. Cross-immunoreactivity of T. glis H3 and protamine to those of primates suggests the evolutionary resemblance of these nuclear basic proteins in primate germ cells. This work was supported by the Thailand Research Fund (Senior Research Fellowship to Prof. Prasert Sobhon).     

The spermatozoa of three species of Xenotrichulidae (Gastrotricha,Chaetonotida): the two “dünne Nebengeisseln” of spermatozoa in Heteroxenotrichula squamosa are peculiar para-acrosomal bodies

The spermatozoa of xenotrichulid gastrotrichs have been studied with the aim of supplying further characters for the phylogenetic analysis of Gastrotricha and to assess the reported biflagellarity of Heteroxenotrichula squamosa. Three species have been examined, belonging to the two hermaphroditic genera of xenotrichulids. The spermatozoa are filiform cells characterized by a scarcely condensed nucleus followed by a single mitochondrion and a flagellum with large accessory fibers. These show an obliquely striated cortex and a core containing some dense material. In Heteroxenotrichula squamosa and Xenotrichula punctata there is also a simple acrosome flanked by two para-acrosomal bodies which are curious long extracellular structures formed by a pile of electron-dense disks connected by thin threads. Xenotrichula intermedia lacks both acrosome and paraacrosomal bodies. The sperm model of xenotrichulids is very different from that of the Macrodasyida and Chaetonotida so far studied, thus supporting an isolated position of the family. The oblique striation of the tail's accessory fibers is similar in to the one period and inclination of the strated cylinder of macrodasyid gastrotrichs, thus being the only spermatological character shared by the two gastrotrich taxa.     

Ultrastructure of Spermatozoa in a Freshwater Fairy Shrimp, <Emphasis Type="Italic">Streptocephalus dichotomus</Emphasis> (Crustacea: Anostraca)

Ultrastructure of spermatozoa in fairy shrimp Streptocephalus dichotomus revealed that they are amoeboid type with no acrosome and flagella. Surface topography of the spermatozoon is smooth with occasional pseudopodial projections. Transmission electron micrographs of spermatozoa show organelle and the mitochondria which is not fused to form the so called ‘Nebenkern’. The testicular lumen reveals spermatozoa in varying sizes and shapes.     

Ultrastructural study of spermatogenesis in the free-living marine nematode <Emphasis Type="Italic">Paracyatholaimus pugettensis</Emphasis> Weiser et Hopper, 1967 (Chromadorida: Cyatholaimidae)

Spermatogenesis and the morphology of mature sperm in the free-living chromadorid Paracyatholaimus pugettensis from the Sea of Japan were studied using transmission electron microscopy. In spermatocytes fibrous bodies (FBs) appear; in spermatids, the synthetic apparatus is located in the residual body, whereas the main cell body (MCB) houses the nucleus, mitochondria, and FBs. The nucleus of the spermatid consists of a loose fibrous chromatin that is not surrounded by a nuclear envelope; centrioles lie in the perinuclear cytoplasm. The plasma membrane of the spermatid MCB forms numerous filopodia. Immature spermatozoa from the proximal part of the testis are polygonal cells with a central nucleus. The latter is surrounded by mitochondria and FBs with poorly defined boundaries. The immature spermatozoa bear lamellipodia all along their surface. Mature spermatozoa are polarized cells with an anterior pseudopodium, which is filled with filaments that make up the cytoskeleton; the MCB houses a nucleus that is surrounded by mitochondria and osmiphilic bodies. In many ultrastructural characteristics, the spermatozoa of P. Pugettensis are similar to those of most nematode species studied so far (i.e., they are ameboid, have no acrosome, axoneme, or nuclear envelope). On the other hand, as in other chromadorids, no aberrant membrane organelles were observed during spermatogenesis of P. Pugettensis.Original Russian Text Copyright © 2004 by Biologiya Morya, Zograf, Yushin.     

Expression of<Emphasis Type="Italic"> FoxE</Emphasis> and<Emphasis Type="Italic"> FoxQ</Emphasis> genes in the endostyle of<Emphasis Type="Italic"> Ciona intestinalis</Emphasis>

We have analyzed the expression patterns of two Fox genes, FoxE and FoxQ, in the ascidian Ciona intestinalis. Expression of Ci-FoxE was specific to the endostyle of adults, being prominent in the thyroid-equivalent region of zone 7. Ci-FoxQ was expressed in several endodermal organs of adult ascidians, such as the endostyle, branchial sac and esophagus. In the endostyle, the pattern of Ci-FoxQ expression was similar to that of CiTTF-1, being prominent in the thyroid-equivalent regions of zones 7 and 8. Therefore, these Fox genes may perform thyroid-equivalent functions in the ascidian endostyle.Edited by N. Satoh     

Ultrastructure of spermatids and spermatozoa in three polychaetes with modified biology of reproduction: Autolytus sp., Chitinopoma serrula,and Capitella capitata

Unlike the primitive type of spermatozoon found in most polychaetes, the spermatozoon of Autolytus has a bilateral symmetry with elongated nucleus, and the mitochondria surround the posterior part of the nucleus. A rather large disk-shaped acrosome is situated along one side of the anterior part of the nucleus. From the anterior margin of the distal centriole emerge long striated rootlets, which run along the nuclear envelope to the anterior part of the nucleus. The spermatozoon of Chitinopoma serrula has an elongated, slightly bent nucleus, a thimble-like acrosome apically on the anterior surface of the nucleus, and an elongated middle piece containing 4 rod-like mitochondria developed from spherical mitochondria surrounding the basal part of the tail flagellum. In the spermatozoon of Capitella capitata, both nucleus and middle piece are elongated compared to the primitive type. The large and conical acrosome is placed asymmetrically at the nucleus and consists of an acrosomal vesicle and subacrosomal substance. The greater part of the middle piece forms a collar around the initial part of the tail flagellum. The cytoplasm of the collar contains granular material. One or two small mitochondria lie around the 2 centrioles at the base of the nucleus.

These types of spermatozoa represent early steps in the evolution of modified spermatozoa combined with changed biology of reproduction. The modified spermatozoa are larger than the primitive ones.     

Characterization and expression analysis of <Emphasis Type="Italic">prohibitin</Emphasis> in the testis of Chinese mitten crab <Emphasis Type="Italic">Eriocheir sinensis</Emphasis>

Prohibitin plays a key role in maintaining mitochondrial membrane integrity and retaining its normal function. We have initially cloned and sequenced the cDNA of prohibitin from testis of the crab Eriocheir sinensis. The 1,357 bp Prohibitin cDNA comprises a 105 bp 5′ untranslated region, a 427 bp 3′ untranslated region and a 825 bp open reading frame. Protein alignment substantiates that the Prohibitin has 70.2, 69.8, 70.5, 70.9, 72.4, 70.6 and 74.9% identity with its homologues in Mus musculus, Homo sapiens, Gallus gallus, Danio rerio, Xenopus tropicalis, Drosophila mojavensis and Aedes aegypti, respectively. In situ hybridization revealed that the Prohibitin mRNA was mainly localized around the proacrosomal vesicle and nucleus membrane in early-stage spermatid. In the following middle stage, Prohibitin mRNA was situated inside the invaginated region of half-moon-like nucleus and surrounded the proacrosomal vesicle. In late-stage spermatid, the mRNA was aggregated in the acrosomal tubule, the band between the acrosome and cup-like nucleus, remanent cytoplasm as well. In the mature sperm, mRNA was only found in the acrosomal tubule and the limited space between the nucleus and acrosome. Therefore, we presume that Prohibitin may fulfill critical functions in the spermiogenesis of Eriocheir sinensis.     

Ultrastructure of the spermatozoon of <Emphasis Type="Italic">Talpa romana</Emphasis> (Mammalia,Lipotyphla)

In this study, we used SEM and TEM to investigate the ultrastructure of spermatozoa from the cauda epididymis of Talpa romana. For comparison, we also analysed spermatozoa from the cauda epididymis of T. europaea captured in the same area. The male gamete of T. romana has a flattened head with an elliptic profile, consisting of a large acrosome and a nuclear region separated by a thin subacrosomal space. At the tip of the nucleus, the subacrosomal space ends in a finger-shaped projection. The tail includes a connecting piece, middle piece, principal piece and end piece. The male gametes of T. romana are substantially similar to those of T. europaea. A comparison with other species of insectivores permits extension of the similarity of sperm features to Scalopus aquaticus and Condylura cristata. Many spermatozoa from the cauda epididymis of T. romana and T. europaea have the tail bent at the annulus, and this is always associated with remnants of cytoplasmic droplets. This morphology is considered to be a common phenomenon.