Comparative oogenesis in cardinal fishes (Apogonidae,Perciformes), with special focus on the adaptive structures of the egg envelopes

The present study compares gonad structure and oogenesis in 30 species of cardinal fish (Apogonidae) from the Indo-Pacific region. In all of the species studied the ovaries are bilobed, with each lobe possessing ovigerous lamellae whose numbers and dimensions increase concomitantly with growth of the fish and reproductive stage, and correlate with the total dimensions of the particular species. In species of the genus Siphamia, the ovarian lobes are associated with a fat body that is not found in species of the other genera studied. Some of the species studied (Siphamia spp.; Archamia spp.) demonstrate group-synchronized maturation of eggs, possibly with only one spawn per reproduction season. However, most of the Apogon spp., and other genera, have unsynchronized cycles, a characteristic of multiple spawners. The various stages of previtellogenic and vitellogenic development are similar in the different species, but differ in timing and dimensions of the oocytes around which the morphogenesis of the egg envelope (chorion) and follicle begins. Differentiation of the egg envelope and follicle cells parallels the vitellogenic stages of egg development. The number of eggs was found to correlate positively with the length of the body and ovary of the fish, whereas egg diameter was found to correlate negatively with body length. The surface of the egg envelope bears ridges that form patterns of various types, whose structures and dimensions seem to be species-specific, and which in most species converge upon the single micropyle on the animal pole. The exposed ridge mazes include a special fibrous web that anchors in the micropyle. The production and importance of these ridges, as well as the correlations between egg numbers per spawn, egg dimensions, and the oral dimensions of the paternal male, are discussed.     

Phylogenetic significance of egg shell variation in tardigrades

Freshwater and semiterrestrial tardigrades produce either smooth shelled eggs laid in the exuvium or ornamented eggs laid freely, in contrast to the more primitive marine heterotardigrades which have smooth shelled free-laid eggs. Data are presented on the observation of free-laid ornamented eggs in the primitive non marine heterolardigrade genus Oreella and in a number ot e.utardigrade genera including Eohypsibius, Amphilus, Minibiotus and Macrobiotus. These findings have provided a sound basis for the formulation of a hypothesis in which the evolution of the tardigrade egg can be explained by two separate events. The first is the acquisition of ornamentation and the second is the utilization of the exuvium for oviposition with subsequent loss of egg ornamentation. This hypothesis is consistent with new taxonomic arrangements within eutardigrade families and genera (based on character) of the animals) which have led to a more uniform assortment in terms of egg types. This means that egg type (smooth or ornamented) becomes a valid taxonomic character at the generic level.     

Closure of the micropyle during embryonic development of some pelagic fish eggs

The fine structure of the egg envelope and micropyle was studied in unfertilized and developing eggs of the flounder Paralichthys olivaceus (Temminck & Schlegel), the Alaska pollack Theragra chalcogramma (Pallas), the Japanese tilefish Branchiostegus japonicus (Houttuyn) and the porgy Pagrus major (Temminck & Schlegel). The outer envelope surface of the unfertilized egg was wrinkled, while the inner surface was folded. The micropyle of the unfertilized egg consisted of a shallow vestibule and a distinct canal. The micropylar region of the inner surface of the envelope had a conical- or bowl-shaped protrusion. In developing eggs, the thickness of the envelope decreased and showed smooth outer and inner surfaces which indicated that it had been stretched tangentially at the time of the perivitelline space formation. The lumen of the micropylar canal was invariably occupied with envelope material. We postulate that the blockage of the micropylar canal is a result of the stretching of the envelope. The closure of the micropyle inhibits sperm and external pathogens from penetrating into the perivitelline space and seems to be involved in both the permanent prevention of polyspermy and the protection of the developing embryo from bacterial infection.     

Three new species of Huffmanela Moravec, 1987 (Nematoda: Trichosomoididae) from the gills of marine fish off New Caledonia

Three new species of Huffmanela, here described from eggs only, are reported from the gills of marine fish caught off Nouméa, New Caledonia. Eggs of Huffmanela branchialis n. sp., from Nemipterus furcosus (Nemipteridae), are 45-52 (mean 48) microm in length and 23-30 (mean 25) microm in width, with thin shells. Each egg is enclosed in a thin membrane forming a spindle-shaped envelope 53-85 (mean 63) microm in length. Eggs of H. filamentosa n. sp., from Gymnocranius grandoculis (Lethrinidae), are 48-53 (mean 50) microm in length and 25-30 (mean 27) microm in width, with thin shells. Each egg bears a few long (150 microm), thin filaments. Eggs of these two new species were compared to those of H. paronai Moravec & Garibaldi, 2000, which are redescribed. Eggs of H. ossicola n. sp. were found within the branchial arch bone of Bodianus loxozonus (Labridae) and also filled the spinal chord bone and other bones. This is the first species of Huffmanela reported from bone tissue. Eggs are large, 72-88 (mean 79) microm in length and 32-40 (mean 36) microm in width, with a very thick shell. Each egg is covered with numerous filaments enclosed in a thin envelope. Fresh eggs were unembryonated, but embryos were visible after incubation in seawater. The three new species can be distinguished from other species of Huffmanela by size and the nature of the egg covering. Egg morphology of and their location in the host suggest different life-cycles: those of the first two species (small eggs, thin shells, egg covering possibly favouring flotation) are released from the gill mucosa with the turnover of living tissues and immediately continue their life-cycle, but eggs of H. ossicola (large eggs, thick shell) are only available for the continuation of the life-cycle after the host's death.     

Fine structure of the egg envelope and the supporting stalk in the dipliran Campodea (Apterygota : Campodeidae)

The eggs and supporting stalks of one Campodea (Apterygota : Campodeidae) species were studied by scanning and transmission electron microscopes. Each egg batch is attached to the substrate by a supporting stalk. The stalk is composed of tangled lamellae and dense annular structures. Both elements are strongly PAS-positive. The eggs are covered by one envelope only. It is built of fine granular, PAS-negative material. The surface of the envelope is smooth and possesses 2–4 ring-like structures, which are likely involved in the attachment of the eggs within a batch.     

Ornamentation of the testa inSedum (Crassulaceae)

A SEM investigation of the seeds of 40 European and 5 Asiatic species ofSedum revealed three fundamentally different types of ornamentation of the testa. The most prominent difference in the ontogeny of the seeds differing in ornamentation is the ovular envelope, which is a mucous membrane enclosing the ovules and young seeds and occurring only in plants whose seeds have a simple type of ornamentation. The ornamentation of the testa was found to be a very constant character. Because the ornamentation is highly correlated with the hybridization pattern of the species it is considered to be an important character in the infrageneric classification ofSedum.     

Effect of population bottlenecks on the egg morphology of introduced birds in New Zealand

Hatching failure is inversely correlated with population bottleneck size among exotic birds introduced to New Zealand, but the mechanism for this is unclear. We assess whether the bottlenecks these species experienced during their introduction have affected hatching failure through deleterious changes in egg morphology. We measured egg size and shape of 13 species that passed through bottlenecks of 11–800 individuals during their establishment in the 19th century. Eggs were also measured in the source populations ('pre-bottleneck') of each species to compare pre- and post-bottleneck egg morphology directly. Significant changes in egg volume were found in six of 13 species, with most laying smaller eggs in New Zealand. Egg shape changed in four of 13 species but there was no directional bias; two species developed more elongated eggs and two species broader eggs. There was no relationship between bottleneck size and change in egg volume, but species passing through severe bottlenecks had greater variability in egg volume and were more likely to have eggs that deviated in shape from their source populations. There was no relationship between changes in either egg volume or shape and rates of hatching failure. Further work is needed to assess whether changes in egg morphology have negative consequences on offspring fitness and whether the observed changes are the result of differing environmental conditions in the introduced range.     

Identification of a novel C1q family member in color crucian carp (Carassius auratus) ovary

Potential roles of C1q/tumor necrosis factor (TNF) superfamily proteins have been observed in vertebrate oogenesis and oocyte maturation, but no ovary-specific member has been identified so far. In this study, we have cloned and identified a novel member of C1q family with a C1q domain in the C-terminal from fully grown oocyte cDNA library of color crucian carp and demonstrated that the gene might be specifically expressed in ovary and therefore designated as Carassius auratus ovary-specific C1q-like factor, CaOC1q-like factor. It encodes a 213 amino acid protein with a 17 amino acid signal peptide. There is only one protein band of about 24.5 kDa in the extracts from phase I to phase IV oocytes, but two positive protein bands are detected in the extracts of mature eggs and fertilized eggs. Furthermore, the mobility shift of the smaller target protein band cannot be eliminated by phosphatase treatment, but the larger protein band increases its mobility on the gel after phosphatase treatment, suggesting that the larger protein might be a phosphorylated form. Immunofluorescence localization indicates that the CaOC1q-like proteins localize in cytoplasm, cytoplasm membrane and egg envelope of the oocytes at cortical granule stage and vitellogenesis stage, whereas they were compressed to cytoplasm margin in ovulated mature eggs and discharged into perivitelline space between cytoplasm membrane and egg envelope after egg fertilization. Further studies on distribution and translocation mechanism of the CaOC1q-like factor will be benefit to elucidate the unique function in oogenesis, oocyte maturation and egg fertilization.     

Dinosaurs: a field guide/The Princeton field guide to dinosaurs

Eggshells from the three extant crocodilian species Crocodylus mindorensis (Philippine Crocodile), Paleosuchus palpebrosus (Cuvier's Smooth-fronted Caiman or Musky Caiman) and Alligator mississippiensis (American Alligator or Common Alligator) were prepared for thin section and scanning electron microscope analyses and are described in order to improve the knowledge on crocodilian eggs anatomy and microstructure, and to find new apomorphies that can be used for identification. Both extant and fossil crocodilian eggs present an ornamentation that vary as anastomo-, ramo- or the here newly described rugosocavate type. The angusticaniculate pore system is a shared character for Crocodylomorpha eggshells and some dinosaurian and avian groups. Previously reported signs of incubated crocodilian eggs were found also on our only fertilised and hatched egg. Paleosuchus palpebrosus presents unique organization and morphology of the three eggshell layers, with a relatively thin middle layer characterised by dense and compact tabular microstructure.     

Cluster of genes encoding the major egg envelope protein of zebrafish

The proteins of fish egg envelopes are encoded by genes that are closely related to the genes for human zona pellucida proteins. A cluster of three genes coding for an egg envelope protein was isolated from the zebrafish, Danio rerio. The three genes, zp2a, zp2b, and zp2c, are located within an 11 kb region and are each comprised of eight exons spanning 1.85 kb. The exon-intron structures of the genes are nearly identical; however, their deduced amino acid sequences diverge at exon 7 (zp2b and zp2c from zp2a) and exon 8 (zp2c from zp2b). Exons 2-7 have a structural organization similar to exons in the carboxy-terminal half of the human zona pellucida ZP1, ZP2, and ZPB genes, suggesting they arose from a common ancestral gene. Sequence comparisons indicate that the deduced zebrafish proteins are most closely related to human ZPB. Zebrafish mRNAs coding for each of the three ZP2 variants have been found either as full-length cDNAs or expressed sequence tags. Distinct from the wf(female) gene of winter flounder which we first reported (Lyons et al., 1993: J Biol Chem 268:21351-21358), expression of the zebrafish zp2 genes was found to be ovary-specific, instead of liver-specific, and the promoter regions of zp2a and zp2b, while different, both contained E-box sequences (CANNTG) that have been demonstrated to be essential for coordination of zona pellucida gene expression in mammalian oocytes. Mixed peptide sequence analysis was used to identify the major polypeptide component of isolated zebrafish egg envelopes as the zp2 gene product.     

Soluble cytokeratins in Xenopus laevis oocytes and eggs

Xenopus oocytes contain a radial network of cytokeratins which seems to fragment during meiosis reinitiation (maturation). The mature egg contains only a cortical network of cytokeratins. We have looked for the presence of soluble cytokeratins in oocytes and unfertilized eggs and have found them in both cases. However, the proportion of soluble to insoluble cytokeratins is slightly higher in the egg than in the oocyte. Soluble cytokeratins incorporate 35S-methionine at a high rate in the oocyte but to a lesser extent in the egg. This suggests that they are biosynthetic intermediates in the oocyte. In the egg, at least a fraction of the soluble cytokeratins may arise from the fragmentation of the polymer which seems to occur during the maturation process. Insoluble cytokeratins are strongly labeled with 32P both in oocytes and eggs. On the other hand only the soluble keratins of the egg incorporate 32P. Since the isoelectric point of soluble and insoluble cytokeratins is the same in oocytes and eggs, their absolute level of phosphorylation probably remains relatively constant. This suggests that: i) phosphate turnover is very slow in oocyte soluble cytokeratins, ii) phosphorylation is not a major way of changing the structural state of cytokeratins in amphibian oocytes and eggs.     

Adhesiveness and surface patterns of eggs in neotropical freshwater teleosts

Eggs of 23 Characiformes and eight Siluriformes, belonging to nine families with diverse reproductive behaviour, were ultrastructurally analysed. The migratory species exhibited non-adhesive eggs, whereas, most of the sedentary species presented some degree of egg adhesiveness. Among the Characiformes, non-adhesive eggs showed zona radiata with pore-canals or a fibrillar net at the surface; weakly adhesive eggs presented only zona radiata with pore-canals while adhesive eggs exhibited zona radiata with apparatus like globules, filaments, villi or honeycomb-like pores depending on the systematic group. The 'jelly' coat is strongly related to the Siluriformes eggs apparently without relationship with adhesiveness. A micropylar disc was present in adhesive eggs of a few species of both Characiformes and Siluriformes. Some patterns were characteristic of the animal pole, others of the vegetal pole, and others were common to both poles. The radial ridges converging to the micropyle in Astyanax bimaculatus lacustris appear to be related to fertilization. In general, egg surface structures in the Characiformes varied according to the genus, whereas all Siluriformes showed a similar egg surface pattern, regardless of the group analysed. Multivariate analysis allowed the identification of eight clusters among the Characiformes and three among the Siluriformes showing relationships between reproductive style and egg characteristics. It is suggested that egg surface and adhesiveness may be related to reproductive patterns and to phylogenesis.     

Oogenesis and ovarian morphology in pollinating and non-pollinating fig wasps: evidence from adult and immature stages

Hymenopteran insects have meroistic polytrophic ovaries characterised by trophocytes associated with oocytes inside the follicles. In pro-ovigenic species, all oocytes mature before emergence and no trace of oogenesis is visible in adult females. Pro-ovigeny is a rare condition among Hymenoptera, but common in pollinating fig wasps. In the present investigation, we studied adult and pupa females of three fig wasp species with different trophic strategies. We demonstrated that females of Pegoscapus aerumnosus and Idarnes spp. have an unusual ovarian organisation (i.e. each ovariole has only one mature egg and no oocyte) that has led to misleading interpretation of fig wasp reproductive anatomy. The ovaries of these studied species have several ovarioles, recognisable by the presence of nuclei of tunica propria cells surrounding them. Each adult wasp ovariole had one mature egg. None of the pupae had mature eggs, but all of them had follicles with oocytes in different developmental stages. The studied fig wasps are pro-ovigenic, irrespective of their trophic strategy, since there were no signs of ovigeny in adult females. We discuss ecological and phylogenetic factors that could play a role in fig wasps reproductive strategies.     

Reproductive cycling and fecundity estimation in the upland bully

The upland bully is a prolific and widespread member of New Zealand freshwater fish communities that may mature at age 1 (or earlier). It begins spawning in spring, laying as many as eight clutches of eggs over the spring and summer, at periods as brief as 13 days. Ovaries contain two modes of maturing oocytes, so that oogenesis must be continual during the spawning season. This poses difficulties in fecundity estimation, which were solved by captive rearing. The ovoid eggs are about 2 mm long; egg size declines with clutch number during the summer, but increases with fish size. Clutch size varied, in 70 spawnings, from 121 to 880 eggs, with total seasonal fecundity up to >5000 eggs. The production of multiple clutches greatly increases seasonal fecundity and may explain the success of this species in sometimes unreliable or harsh environments.     

The use of poly-L-lysine to facilitate examination of sperm entry into pelagic, non-adhesive fish eggs

The fish egg is surrounded by a thick envelope called the chorion. The fertilizing spermatozoon enters the egg through a canal-like structure in the chorion, the micropyle. Examination of micropyle at fertilization is difficult if eggs are large and have no distinct landmarks surrounding the micropyle, or if they are positively buoyant in water. Eggs of many commercially important fishes (e.g., flounder, sea bream and eel) are buoyant in water or only slightly adhere to solid objects (e.g., sands, rock and water plants), which makes observation of spermatozoa at fertilization difficult. Here, we report that such eggs can be firmly attached to plastic and glass dishes that have been previously coated with poly-L-lysine. These adhering eggs can be fertilized and develop normally on the dishes. Observations of micropyles of three fish species, before and after sperm entry are presented.     

Electron microscopic observations on sperm entry and pronuclear formation in naked eggs of the rose bitterling in polyspermic fertilization

Unfertilized eggs of the rose bitterling (Rhodeus ocellatus ocellatus) were squeezed out of females that had an elongated ovipositor and were dechorionated mechanically with fine forceps in physiological saline. The dechorionated eggs were transferred into fresh water then inseminated at once by spermatozoa of the same species. A large number of spermatozoa was found on the surface of eggs that had not yet had cortical reaction following insemination. The surface of the naked eggs responded by formation of many small cytoplasmic protrusions (viz., fertilization cones) at sperm attachment sites. The formed fertilization cones were rosettelike structures formed by the aggregation of some bleblike swellings devoid of microvilli and microplicae. About 10 min after insemination, the fertilization cones retracted, but marks of their presence characterized by less microvilli and microplicae remained in the eggs 15 min after insemination. Many spermatozoa penetrated into the cytoplasm of each naked egg. The sperm nuclear envelope disappeared by means of vesiculation resulting from fusion of the inner and outer membranes. The sperm nucleus decondensed and developed into a larger male pronucleus. Smooth-surfaced vesicles surrounded the decondensing sperm nucleus and formed the new male pronuclear envelope. Sperm mitochondria and flagella were found in the egg 15 min after insemination. The response of the egg surface to sperm entry and pronucleus formation are discussed.     

Ultrastructure of the Secondary Egg Envelope of Cyprinodontidae of the Genus Epiplatys Gill, 1862 (Pisces,Teleostei)

Abstract The structure of the secondary egg envelope of seven species of Cyprinodontidae belonging to the genus Epiplatys was observed by transmission and scanning electron microscopy. The transverse structure of the secondary envelope examined by transmission electron microscopy is a constant feature in all the species studied and represents a good criterion for identification. In contrast, the surface structure of the secondary envelope when observed by scanning electron microscopy exhibits a great diversity of form. It is possible to distinguish each of the seven different species by the pattern of ornamentation, the width of the polygons and the size of the granules comprising them on the secondary egg envelope.     

前寒武纪瓮安生物群具围卵腔结构的胚胎及其发育序列

通过对贵州瓮安新元古代陡山沱组瓮安微型球状化石SEM的观察,发现代表未分裂卵,和2、4、8、16不同分裂阶段的胚胎。这些未分裂的卵和处于不同发育阶段的胚胎化石不仅大小和卵壳表面装饰相同,而且它们均具明显的围卵腔和球形卵裂等特征,清楚表明这些卵和胚胎均为同一物种的产物。由于这些卵的表面装饰与现生某些甲壳类休眠卵极其相似,曾被解释为动物的休眠卵;当前不同卵裂阶段胚胎的发现表明它们不是休眠卵,而是处于发育过程的卵。围卵腔不仅是现生两侧对称动物早期胚胎的常见构造,而且具围卵腔构造的未分裂卵和不同卵裂阶段的胚胎化石在瓮安动物群也十分常见。除了以上具表面装饰胚胎化石的大量发现外,文中还报道围卵腔构造在表面光滑胚胎化石中的发现。这些具围卵腔结构的胚胎化石主要为两侧对称动物的胚胎。     

Voltage response to fertilization and polyspermy in sea urchin eggs and oocytes

Successful collision rates of sperm with eggs and oocytes of the sea urchins Psammechinus microtuberculatus and Paracentrotus lividus have been studied using an electrophysiological method. A monospermic response in eggs consists of a 1- to 2-mV step depolarization of the egg plasma membrane accompanied by an increase in voltage noise. The step precedes the main positive-going depolarization by approximately 13 sec at room temperature. If other successful collisions occur during this 13-sec period (indicated by additional steps), the egg is polyspermic. It is shown by direct observation that each step depolarization signifies the entry of a single sperm. No evidence for an electrically mediated fast block was found. The average rate of successful sperm-egg encounters increases with sperm density, although individual steps appear to occur randomly. Step depolarizations also occur in oocytes, however, they usually decay after several seconds and are not followed by a large, positive-going depolarization. The rate of occurrence of such steps increases with sperm density over the range 10⁵ to 10⁹ sperm/ml. The original evidence of Rothschild and Swann for a fast partial block is compared with a model of polyspermy suggested by our experiments. Reasonable agreement between our method of counting successful collisions (in oocytes and eggs) and the method used by Rothschild and Swann (for eggs) was obtained for sperm densities below 10⁶/ml. The results diverge for higher sperm densities, our method giving higher values. A test for the hypothesis of a fast partial block to polyspermy is suggested, using our method of counting successful sperm-egg collisions.     

Variation in ovariole number and egg size of species of Dacus (Diptera; Tephritidae) and their relation to host specialization

Abstract. 1. An analysis of the number of ovarioles/ovary is presented for fourteen species of Dacus ranging from broad generalists (D.tryoni (Frogg.)) to strict host specialists (D.aglaia Hardy). For eight species data on egg size and the incubation time of eggs is also presented. Interrelationships among these life history variables and their relationships to host specialization were examined. 2. There was a broad positive relationship between the breadth of the natural host range (i.e. excluding cultivated fruits) and ovariole number (and hence potential fecundity). Highly polyphagous species had thirty-five to forty ovarioles/ovary while the specialists ranged from eight to twenty. 3. D.musae (Tryon), a specialist on banana, was an exception having an ovariole number similar to that of the most polyphagous species. 4. There was no clear relationship between egg size and ovariole number across eight species, though monophages were not fully represented in this analysis. Among these eight species there was no significant relationship between wing length and egg length, nor was there any such relationship among individuals within species. 5. There was considerable variation among species in the incubation time (at 25°C) for eggs, ranging from 24 h for D.cucumis French to 53 h for D.cacuminatus (Hering). This variation was only weakly related to egg size, though D.cucumis produced the largest eggs of any species examined. 6. Possible explanations for a relationship between potential fecundity and host range are discussed in terms of the balance between the abundance of host resources for generalists and their predictability for specialists. However, the observed relationship between ovariole number and host range must remain tentative, given that only fourteen species from one genus have been examined. Considerably more data is required for other dacines, for species in other tephritid subfamilies, and for other phytophagous insect groups to determine whether the relationship is generally applicable.