Ontogeny of the collar cord: Neurulation in the hemichordate Saccoglossus kowalevskii

The chordate body plan is characterized by a central notochord, a pharynx perforated by gill pores, and a dorsal central nervous system. Despite progress in recent years, the evolutionary origin of each of theses characters remains controversial. In the case of the nervous system, two contradictory hypotheses exist. In the first, the chordate nervous system is derived directly from a diffuse nerve net; whereas, the second proposes that a centralized nervous system is found in hemichordates and, therefore, predates chordate evolution. Here, we document the ontogeny of the collar cord of the enteropneust Saccoglossus kowalevskii using transmission electron microscopy and 3D‐reconstruction based on completely serially sectioned stages. We demonstrate that the collar cord develops from a middorsal neural plate that is closed in a posterior to anterior direction. Transversely oriented ependymal cells possessing myofilaments mediate this morphogenetic process and surround the remnants of the neural canal in juveniles. A mid‐dorsal glandular complex is present in the collar. The collar cord in juveniles is clearly separated into a dorsal saddle‐like region of somata and a ventral neuropil. We characterize two cell types in the somata region, giant neurons and ependymal cells. Giant neurons connect via a peculiar cell junction that seems to function in intercellular communication. Synaptic junctions containing different vesicle types are present in the neuropil. These findings support the hypotheses that the collar cord constitutes a centralized element of the nervous system and that the morphogenetic process in the ontogeny of the collar cord is homologous to neurulation in chordates. Moreover, we suggest that these similarities are indicative of a close phylogenetic relationship between enteropneusts and chordates. J. Morphol., 2010. ©2010 Wiley‐Liss, Inc.     

The spermatozoon of the Echiniscidae (Tardigrada,Heterotardigrada) and its phylogenetic significance

The spermatozoon ultrastructure of four species of moss-dwelling Heterotardigrada belonging to four genera of Echiniscidae, namely Pseudechiniscus juanitae, Echiniscus duboisi, Novechiniscus armadilloides and Antechiniscus parvisentus, was investigated. In all species, the testicular male gamete is similar in morphology and in length. The spermatozoon is made up of a long head, consisting of a cylindrical acrosome and an oval or rod-shaped nuclear region which contains a nucleus with osmiophilic and electron-dense chromatin, and a tapering tail, with a "9+2" axoneme. An elongated sack-like structure originates from the posterior part of the head, extending beyond the main axis of the cell and running parallel to the tail. It consists of two parallel tubular regions which sometimes form a strict double helix and contain two voluminous, "free" mitochondria with unmodified cristae. In addition, a voluminous vesicle is present laterally to the centriole or between the end of the nucleus and the beginning of the mitochondria, limited by two cytomembranes and filled with electron-lucent and granular material. The male gametes representative of these moss-dwelling Echiniscidae are very similar to the spermatozoa of the marine Echiniscoididae Echiniscoides sigismundi. This close similarity emphasises that habitat changes have had little influence on the organisation of the sperm cell representative of Echiniscoidea. Spermatozoon characters which could be useful for phylogenetic studies on Tardigrada are discussed.     

Structure,Ultrastructure, and Function of the Preoral Heart-Kidney in Saccoglossus kowalevskii (Hemichordata,Enteropneusta) Including New Data on the Stomochord

The heart-kidney of Saccoglossus kowalevskii, which is situated within the anterior preoral proboscis coelom (protocoel), consists of the stomochord, pericardium, heart sinus, and glomerulus. The stomochord, a diverticulum of the gut, is characterized by vacuolated epithelial cells surrounded by basal lamina and connective tissue. The pericardium, a myoepithelium, lies dorsal to the central heart sinus. Opening into the protocoel and connecting with the outside via the proboscis pore is the protocoel duct, which is, in part, composed of multiciliated absorptive epithelial cells. Perfusion of the dorsal trunk vessel with vital dyes reveals a rapid flow of blood into the glomerular blood vessels. Examination of the permeability characteristics of the extracellular matrix underlying the glomerular podocytes reveals the movement of iron dextran (mol. wt 5000 daltons) from the central heart sinus into the protocoel. Iron dextran uptake by glomerular cells and protocoel lining cells is demonstrated. These results suggest that vascular fluid is filtered by the glomerulus, producing a primary urine in the protocoel which may be modified as it passes over the peritoneum, through the protocoel duct, and out of the proboscis pore. New data concerning the morphology of the stomochord are presented. The controversial homology between the hemichordate stomochord and the chordate notochord is addressed.     

The gene for a cytoplasmic intermediate filament (IF) protein of the hemichordate Saccoglossus kowalevskii; definition of the unique features of chordate IF proteins

We have isolated full length cDNAs encoding a cytoplasmic intermediate filament (IF) protein of a priapulid (Priapulus caudatus) and of a hemichordate (Saccoglossus kowalevskii) and determined the organisation of the hemichordate gene. As expected, the priapulid protein shows the long coil 1b subdomain and the lamin tail homology segment already known for cytoplasmic IF proteins from 11 other protostomic phyla. Surprisingly, the hemichordate protein follows in domain organisation much more closely the protostomic IF proteins than the chordate IF proteins. Thus, the lack of a lamin tail homology segment together with a deletion of 42 residues in the coil 1b domain is a molecular feature restricted to the chordates. We propose that the known IF subfamilies I to IV developed by gene duplications and sequence drift after the deletion in coil 1b arose at the base of the chordate branch.     

The postanal tail of the enteropneust Saccoglossus kowalevskii is a ciliary creeping organ without distinct similarities to the chordate tail

Stach, T. and Kaul, S. 2011. The postanal tail of the enteropneust Saccoglossus kowalevskii is a ciliary creeping organ without distinct similarities to the chordate tail. —Acta Zoologica (Stockholm) 92 : 150–160. The postanal tail of chordates is one of the key characters in chordate evolution and it has been suggested to be homologous to the postanal tail of harrimaniid enteropneusts. We present electron microscopic data of the ontogeny of the postanal tail in the enteropneust Saccoglossus kowalevskii. The postanal tail develops as a ventral posterior allometric outgrowth with a ventral extension of the telotroch. Transmission electron microscopy of serial sections reveals the epidermal organization of the postanal tail with the exception of short, bilaterally symmetric extensions of the paired metacoels. The epidermis cells are connected by apical junctions, rest basally on the extracellular matrix surrounding the mesoderm, and possess a basiepidermal nerve net. The ventral cells in the postanal tail are multiciliated and used for creeping. Dorsal cells are monociliated with numerous microvilli. Two types of glandular cells are present among the epidermis cells. The mesoderm cells contain myofilaments. We were unable to detect anatomical structures similar to the ones present in the postanal locomotory tail of chordates, such as notochord, neural tube, or endodermal strand. Thus, results of our anatomical study do not support homology of the postanal chordate tail and the postanal tail of harrimaniid enteropneusts.     

Ultrastructure of the spermatozoon of Lepidogalaxias salamandroides and its phylogenetic significance

The spermatozoon of Lepidogalaxias salamandroides possesses an acrosome (putative), one or two perforatoria (putative) but no nine-triplet centrioles. Two elongated mitochondria (12 μm long) are situated in parallel between the nucleus (20 μm long) and the axoneme (53 μm long). The above features are unique among other teleosts with internal fertilization. The presence of an “acrosome” in this primitive teleost supports the hypothesis that this structure has been secondarily lost in teleosts during evolution. The uncertainty of phylogenetic placement of this fish is reflected by its unique sperm ultrastructure.     

The ultrastructure of the spermatozoon of Haplotaxis ornamentus (Annelida,Oligochaeta, Haplotaxidae) and its phylogenetic significance

Summary The spermatozoon of Haplotaxis ornamentus has characteristics common to all oligochaete sperm: filiform; primary acrosome vesicle carried on an acrosome tube and containing an axial rod (perforatorium) in an invagination (subvesicular space or secondary acrosomal invagination); an elongate, highly condensed cylindrical nucleus followed by a cylindrical midpiece of radially adpressed mitochondria not penetrated by the axoneme; a single (distal) centriole persistent, though modified, at maturity; axoneme with 9 doublets, each with two outer glycogen granules, and centrally two singlets accompanied by two solid fibres. A peculiar haplotaxid combination of characters (none unique) is slight withdrawal of the primary vesicle into the acrosome tube with a strongly emergent capitulate axial rod and moderately short midpiece. This ultrastructure is consistent with location of the Haplotaxidae at the base of the Haplotaxida (Haplotaxina — Alluroidina — Moniligastrina — Lumbricina). Tubificida sperm, although also plesiomorph for the Oligochaeta, have the autapomorphy elongate periaxial sheath (secondary tube), excepting the Phreodrilidae whose sperm show convergent resemblances to the Lumbricina. The term annuloid has been introduced for annulus-like structures of varied origins.     

Giant spermatozoon coiled in small egg: fertilization mechanisms and their implications for evolutionary studies on Ostracoda (Crustacea)

Ostracods of the superfamily Cypridoidea have giant spermatozoa. However, little data exist on the sperm-egg interaction in this group: only two publications have so far given the most ambiguous indication that the entire sperm enters the egg on fertilization. These assumptions have not yet been tested with modern techniques, nor has their putative value for developmental and evolutionary investigations been realized. The present paper gives the first, clear, light- and scanning electron microscopical evidence of the entire giant ostracod spermatozoon being incorporated into the egg. Coiling of the sperm underneath the egg shell is shown in the early zygotes of the species Mytilocypris praenuncia and Pseudocandona marchica. Additionally, data on the morphology of female and male reproductive tracts are given for M. praenuncia. Hypotheses on the evolution of giant filiform sperm in the Animal Kingdom are reviewed, and their applicability to ostracods is discussed. The demonstrated ingression of the entire sperm implies the entry of the two giant paternal mitochondrial derivates into the zygote in Cypridoidea and potentially casts doubt upon the dogma of strict maternal inheritance of mitochondrial DNA. Evidence of paternal inheritance of mtDNA in several organisms has recently given rise to a controversial debate on this issue; the possible significance of this phenomenon for molecular studies on ostracod phylogeny and evolution is discussed.     

The ultrastructure of the spermatozoon of Neochasmus sp. (Cryptogonimidae, Digenea, Trematoda) and its phylogenetic significance

Jamieson B. G. M. and Daddow L. M. 1979. The ultrastructure of the spermatozoon of Neochasmus sp. (Cryptogonimidae, Digenea, Trematoda) and its phylogenetic significance. International Journal for Parasitology12: 547–559. The typical spermatozoon is filiform, monopartite and internally strongly asymmetrical, with a mean length of 254 μm. Its two axonemes are wholly incorporated though not deeply. The six major regions, in anteroposterior (proximal-distal) sequence, are: acrosomoid, prenuclear, uniaxonemal, nuclear, intermediate (amitochondrial), posterior mitochondrial and posterior amitochondrial regions. Centrioles and flagellar rootlets, present in the spermatid, are absent from the spermatozoon. The right axoneme (axial unit) originates more anteriorly than the left and anteriorly bears an acrosome-like projection (acrosomoid). Spinose regions carrying epiplasmalemmal projections are reported for the first time in the Digenea. Dimorphism exists as replacement of the typical posterior mitochondrial and amitochondrial regions by an amitochondrial region characterized by asymmetrical distribution of spines around one axoneme. Excepting known schistosomatid sperm, the digenean sperm is diagnosed by the combination of three characters: cortical microtubules, two fully incorporated 9 + 1 axonemes and presence of mitochondria although no one of these characters is limited to the group. Subtle intertaxon variations occur.     

Ultrastructure of the spermatozoon of Centroderma spinosissima (Stossich, 1886) (Digenea: Mesometridae) and its phylogenetic potential

The Mesometridae includes only five genera and eight species. The available data on the ultrastructure of sperm cells of mesometrid species referred to two species only, Elstia stossichianum and Wardula capitellata. The present study revealed the ultrastructure of the spermatozoon of a third genus and third species of Mesometridae, Centroderma spinosissima. The mature spermatozoon of C. spinosissima presents two axonemes with different lengths of the Ehlers’ 9 + ‘1’ trepaxonematan pattern, a nucleus, two mitochondria, two bundles of parallel cortical microtubules, external ornamentation of the plasma membrane, a lateral expansion, spine-like bodies, cytoplasmic ornamented buttons and granules of glycogen. The spermatozoon of C. spinosissima is similar to those of the previously studied mesometrids. However, some peculiarities such as the presence of two mitochondria, the disposition of the external ornamentation of the plasma membrane and the morphology of the posterior spermatozoon extremity, characterize the male gamete of C. spinosissima. Moreover, the presence of cytoplasmic ornamented buttons is a characteristic found only in the mature spermatozoon of mesometrids and it probably represents an autapomorphy for this family.     

Sperm-oocyte membrane fusion in the human during monospermic fertilization

Sperm-oocyte membrane fusion has been observed during monospermic fertilization of a human oocyte in vitro. Women were stimulated with both clomiphene citrate and human menopausal gonadotropin and were given human chorionic gonadotropin before a LH-surge. Twelve oocytes, collected at laparoscopy from six women who became pregnant by IVF, were allowed to mature for 7–14 hours in vitro and inseminated with preincubated sperm, fixed between 1–3 hours after insemination, and examined by transmission electron microscopy. Membrane fusion had occurred in one ovum 2 hours after insemination, and the oocyte had resumed maturation and was at anaphase II of meiosis. Cortical granules had been exocytosed, and some of their contents were visible at the surface close to the oolemma all around the oocyte. The sperm that fused with this oocyte was acrosome-reacted and had been partly incorporated into the ooplasm, while the anterior two-thirds of its head was phagocytosed by a tongue of cortical ooplasm. Membrane fusion had occurred between the oolemma and the plasma membrane overlying the postacrosomal segment of the sperm head, posterior to the equatorial vestige. Sperm chromatin had not decondensed, and serial sections revealed a midpiece attached to the basal plate and a tail located deeper in the ooplasm, all devoid of plasma membrane. Supplementary sperm penetrating the inner zona, approaching the perivitelline space, had undergone the acrosome reaction but had a persistent vestige of the equatorial segment of the acrosome with intact plasma membrane. Evidence of sperm chromatin decondensation was seen in other oocytes, 3 hours after insemination, which were at telophase II of meiosis. Eight oocytes penetrated by sperm were monospermic, while four were unfertilized. The general pattern of sperm fusion and incorporation appears to conform to that seen in most other mammals. The study also reveals that sperm have to complete the acrosome reaction before fusing with the egg.     

The antibacterial activity of the hemichordate Saccoglossus ruber (Enteropneusta)

Antibacterial activity of crude whole body homogenates and mucus from the body surface of the hemichordate Saccoglossus ruber was detected using a turbidometric and plate assay. Activity was found to be strongest against marine bacteria and significantly higher against bacteria isolated from the immediate environment of the animals. The whole body nomogenate was less effective at inhibiting growth of the seven terrestrial bacteria tested. The antibacterial factor from the homogenate retained its activity after boiling and was stable at room temperature for at least 8 weeks, suggesting that it was nonproteinaceous. The loss of all activity during dialysis also indicated that a low molecular weight substance was involved. The antibacterial factor from the mucus and homogenate had similar properties, so that the same molecule may have been involved. The possibility that the antibacterial factor may be related to the halogenated indoles and phenols occurring in other enteropneusts is discussed.