Hatching mechanism and delayed hatching of the eggs of three broadcast spawning euphausiid species under laboratory conditions

Three different egg hatching mechanisms were observed underlaboratory conditions in Euphausia pacifica Hansen, Thysanoessaspinifera Holmes and Thysanoessa inspinata Nemoto: backward,forward and flipping. Like all broadcast spawning euphausiids,these species usually hatch as nauplius 1 (N1). Some hours beforehatching the vitelline membrane breaks and the embryo is freelysuspended within the chorion; later the embryo takes on a slightlyoval shape. When ready to hatch, the N1 pushes against the chorionwith the posterior part of the abdomen producing a protuberance.No spine or egg tooth is present to break the chorion. The pressurebreaks the chorion, and the nauplius pushes itself backwardswith the first and second antennae and mandible to slide fromthe chorion. After about three quarters of the body is outside,the nauplius brings all the appendages together to move backwardswithout becoming stuck in the chorion. This is the backwardhatching mechanism. The vitelline membrane remains within theegg after the nauplius leaves the chorion. Hatching takes 5–20s, and most of the eggs in a clutch hatch during <2 h. Severaleggs of E. pacifica hatched as meta-nauplii (MN) (>200 hafter spawning) or as calyptopis 1 (C1) stage (>232 h), ratherthan as N1. Delayed hatching of embryos also was observed inT. spinifera as nauplius 2 (N2) (>120 h) or as MN stage (>180h), and in T. inspinata as N2 (106 h) after spawning. Eggs withlarvae in stages of development beyond N1 have not been observedfrom preserved zooplankton samples. However, eggs spawned inthe field and incubated in the laboratory also had extendeddevelopment and late hatching but with low frequency (<0.06%).It is proposed that, if the backward hatching mechanism fails,alternate hatching mechanisms can be used by the euphausiid.There is high flexibility in their hatching modes. The N2 andMN break the chorion with the first and second antennae, hatchingforwards, and the C1 breaks it with the telson spines and byflipping of the abdomen, resembling the decapod hatching mechanism.Delayed hatching using the forward and flipping mechanisms wereassociated with low hatching success in comparison with thebackward hatching mechanism.     

Structure and enzymatic removal of the chorion of embryos of the Nile tilapia

Dechorionation or partial digestion of the egg chorion is necessary for introducing embryonic stem cells into blastulas for chimera production and for harvesting blastulas. Several methods to digest the Nile tilapia Oreochromis niloticus chorion were tried and it was found that the chorions of most clutches of eggs were digested in <3 h using hatching medium [produced by allowing embryos to hatch in Hanks balanced salt solution (HBSS) in an incubator], or 2 mg ml⁻¹ pronase P6911 in 10% Ca/Mg free HBSS. The chorion of Nile tilapia possesses multiple lamellae as found in most teleost species that have been studied. It was found to be thinner than that of medaka Oryzias latipes and thicker than that of zebrafish Danio rerio . During natural hatching the chorion was digested from the inner surface, and tail movements helped to break the remaining chorion; however, chorion digestion has to be complete for experimental dechorionation, because digestion starts at the external surface. The zona radiata externa remained intact after experimental digestion with hatching media but was disrupted by pronase. Embryos dechorionated at the cleavage or blastula stage only survived for 2 or 3 days, but some dechorionated at the gastrula stage or early segmentation stage developed until the natural hatching time. If the chorion was partially digested at the cleavage or blastula stage, some embryos survived to hatch.     

Morphometric study of embryonic development of Macrobrachium borellii (Arthropoda: Crustacea)

Summary

Embryo morphometry and developing time from just-laid eggs until hatching were described in the palaemonid prawn, Macrobrachium borellii, using a rapid non-invasive staging method. Embryos were kept in the laboratory under controlled conditions and development divided into seven stages according to major morphological characteristics. This lecithotrophic, freshwater shrimp has a highly abbreviated type of hatching development after 39 ± 2 days as postlarvae at 24°C. Morphometry was recorded using a stereoscopic microscope with an image analyzer. Area, perimeter, maximum and minimum diameters and shape were measured in yolk sac, egg-coat and eye, respectively, and they were statistically selected as the best to define the stages. Eggs are ovoid with a maximum diameter that varies from the moment of oviposition to the time of hatching from 1.5 to 2.0 mm, respectively. Water content and egg size increase along with development, whereas egg shape only varies just before hatching when the egg becomes strongly ovoid. Egg coat and eye variables significantly increase as the embryo develops while all yolk variables decrease as the embryo consumes the vitellus. Yolk represents more than 95% of the egg at the time of oviposition, falling to 22% by the time of hatching. The major yolk area decrease is observed between stages 4 and 5, which is coincident with a marked increase in the catabolism. Using only egg coat, yolk and eye shape and maximum diameter, a researcher can straightforwardly identify a developing stage with an accuracy ranging from 70 to 100%. This tool may be employed in other species provided they have transparent chorion.     

Relationships between egg size, chick size at hatching, and chick survival in the Whimbrel Numenius phaeopus

The relationships between egg size, chick size at hatching and chick survival in Whimbrels Numenius phaeopus were studied over a three-year period in the Shetland Isles. Three measurements of chick size at hatching were all positively correlated with egg volume, though the relationship was strongest with hatchling body-weight. In two of the three years the proportion of chicks from a brood which survived to fledging increased significantly with the mean hatching weight of chicks in the brood. Within broods, a significant effect of hatching weight on survival was detectable only up to 7 days after hatching. Between years the egg volumes and hatchling weights of individual female Whimbrels showed relatively little variability, indicating that these attributes could be controlled to a large extent by inheritance.     

Phenotypically plastic responses of green frog embryos to conflicting predation risk

Predators have been shown to alter the timing of switch points between life history stages, but few studies have addressed switch point plasticity in prey exposed simultaneously to conflicting predation pressure. We tested hatching responses of green frog (Rana clamitans) embryos subject to perceived predation risk from chemical cues released by two stage-specific predators, predicting that these predators would elicit: (1) directional hatching responses when presented independently, and (2) intermediate phenotypic responses when presented simultaneously. R. clamitans embryos in outdoor exclosures were exposed to cues from an egg predator (freshwater leeches; Nephelopsis obscura), a larval predator (dragonfly nymphs, Aeschna canadensis), and both predators in a 2 × 2 factorial experiment, and changes in hatchling size, hatchling developmental stage, and hatching time were compared to those for control embryos. Leeches alone induced embryos to hatch at a smaller size and an earlier developmental stage than controls, while dragonfly nymphs elicited a delay in egg hatching time that was associated with larger size and later developmental stage at hatching. Embryos failed to respond to simultaneous exposure to both predators, implying that responses to each occurred concurrently and were therefore dampened. Our results indicate that prey under threat from conflicting predators may manifest intermediate defensive phenotypes. Such intermediate responses may result in elevated rates of prey mortality with possible consequences at the population level.     

Role of epidermal cilia in development of the Australian lungfish,Neoceratodus forsteri (Osteichthyes: Dipnoi)

In common with the embryos of other anamniotes, young of the Australian lungfish, Neoceratodus forsteri, have ciliated cells in the epidermis. These first appear at stage 28, ˜ 10 days before hatching, and develop progressively to a peak in numbers and in activity at stage 44, just after hatching. After this point, ciliary action in the epidermal cells slowly declines, and cilia disappear completely from the outer surface of the hatchling by stage 52. Cilia are lost earlier from the oral epithelium, between stages 45 and 46, and from the epithelium covering the gills and lining the operculum at stage 51, although they are retained in the nares and in the cavity of the olfactory organ. To assess possible functions for the ciliated epidermis in lungfish hatchlings, the presence of cilia in the epidermis of young N. forsteri is compared with landmarks of development. The ciliated epidermal cells are not associated with movements of the embryo within the egg capsule, nor are they a part of a feeding mechanism. They are not related to oxygen uptake. The ciliated epidermis appears to function as a mechanism for clearing the animal of particles and settling organisms before hatching, when the egg membranes have developed holes, and after hatching, when the young fish is living among the submerged rootlets of trees growing on the river bank or in dense stands of aquatic plants. The function of a ciliated epidermis in N. forsteri hatchlings in relation to microhabitat is discussed. © 1996 Wiley-Liss, Inc.     

Hatching enzyme of the ovoviviparous black rockfish Sebastes schlegelii- environmental adaptation of the hatching enzyme and evolutionary aspects of formation of the pseudogene

The hatching enzyme of oviparous euteleostean fishes consists of two metalloproteases: high choriolytic enzyme (HCE) and low choriolytic enzyme (LCE). They cooperatively digest the egg envelope (chorion) at the time of embryo hatching. In the present study, we investigated the hatching of embryos of the ovoviviparous black rockfish Sebastes schlegelii. The chorion-swelling activity, HCE-like activity, was found in the ovarian fluid carrying the embryos immediately before the hatching stage. Two kinds of HCE were partially purified from the fluid, and the relative molecular masses of them matched well with those deduced from two HCE cDNAs, respectively, by MALDI-TOF MS analysis. On the other hand, LCE cDNAs were cloned; however, the ORF was not complete. These results suggest that the hatching enzyme is also present in ovoviviparous fish, but is composed of only HCE, which is different from the situation in other oviparous euteleostean fishes. The expression of the HCE gene was quite weak when compared with that of the other teleostean fishes. Considering that the black rockfish chorion is thin and fragile, such a small amount of enzyme would be enough to digest the chorion. The black rockfish hatching enzyme is considered to be well adapted to the natural hatching environment of black rockfish embryos. In addition, five aberrant spliced LCE cDNAs were cloned. Several nucleotide substitutions were found in the splice site consensus sequences of the LCE gene, suggesting that the products alternatively spliced from the LCE gene are generated by the mutations in intronic regions responsible for splicing.     

Egg development, hatching and early life-history of Dissonus manteri (Copepoda), parasitic on the gills of coral trout, Plectropomus leopardus

The early life-history of Dissonus manteri differs markedly from that of Caligidae and from that of its congener Dissonus nudiventris. The uniseriate egg strings are of typical caligid form, but the stage hatching from the egg is a preecdysis nauplius which moults immediately to the copepodid. The empty naupliar cuticle remains tethered to the egg string by a pair of posterior threads which attach to the internal surface of the egg membrane. The duration of egg development is 112 h and the mean duration of hatching is 5.2 h at 25 degrees C. Aeration and water movement are important for successful hatching. Under these conditions, hatching success is 94.8%, but decreases to 48.4% in static water with no aeration. Mean survival time of the copepodid in the absence of a host is 41.4 h at 25 degrees C. The reduction of early free-living stages in the life-cycle and the high survival rate from egg to copepodid could be important in future coral trout aquaculture, since the potential for re-infection by this parasite is very high.     

Embryonic growth and mobilization of energy and material during incubation in the checkered keelback snake, Xenochrophis piscator

We collected 20 checkered keelback snakes (Xenochrophis piscator) to study embryonic growth and mobilization of energy and material during incubation. Females laid eggs between late May and late June. The eggs were incubated at 27 degrees C (+/-0.3). One egg from each clutch was dissected at five-day intervals starting at oviposition. The mean incubation length at 27 degrees C was 48.9 days. We identified three phases of embryonic growth or yolk depletion in X. piscator. Phase 1, between oviposition and Day 20, was one of minimal transfer of energy and material from yolk to embryo. Phase 2, between Day 20 and Day 39-40, was characterized by increasingly rapid embryonic growth or yolk depletion. Phase 3, between Day 39-40 and hatching, was characterized by reduced embryonic growth or yolk depletion. Approximately 71% of dry mass, 53% of non-polar lipids and 66% of energy were transferred from the egg contents to the hatchling during incubation. Our data confirm that oviposition is not timed to coincide with the onset of rapid embryonic growth in oviparous squamate reptiles. The greater conversion efficiencies of energy and material from egg to hatchling in snakes can be attributed to their lower energetic costs of embryonic development and greater residual yolk sizes.     

Pre- and postnatal energetics of the North Island brown kiwi (Apteryx mantelli)

In four eggs and four chicks of the North Island brown kiwi (Apteryx mantelli) we measured pre- and postnatal oxygen consumption rate (VO(2)) daily from day (d)-75 (prior to hatching) until d+25 (after hatching). The increase of embryonic VO(2) reaches a plateau phase between d-22 and d-5 (0.113 ml O(2)/g/h=59.6% of allometrically expected value of a typical 416-g egg). Mean total O(2) uptake per egg (43.01 l O(2)) corresponds to an energy turnover rate of 2.04 kJ/g during embryonic development. This is nearly identical to the expected value for all birds (2.00+/-0.8 kJ/g). Hence, the kiwi neither 'gained nor lost energy' (Calder, 1979.Biosci. 8, 461-467) by its extreme prolongation of incubation time; it is as efficient as other avian embryos. The kiwi embryo expends only approximately 17% (847 kJ) of the energy originally stored in the egg (4942 kJ). Forty-eight percent of the egg's initial yolk mass can be found as spare yolk in the hatchling and can serve as the chick's sole source of energy and substrate for tissue production for up to at least 17 days after hatching.     

Purification and partial characterization of high choriolytic enzyme (HCE), a component of the hatching enzyme of the teleost, Oryzias latipes

The hatching enzyme is an embryo-secreted enzyme(s) which digests the egg envelope, allowing the embryo to emerge at the time of hatching. The hatching enzyme of the fish, Oryzias latipes, has recently been found to consist of two kinds of proteases which may digest the inner layer of chorion (egg envelope) cooperatively [Yasumasu, S. et al. (1988) Zool. Sci. 5, 191-195]. In the present study, one of them, high choriolytic (egg envelope digesting) enzyme (HCE) was purified and some biochemical and enzymological properties were examined. The enzyme was a basic protein with a molecular weight of about 24 kDa, and exhibited choriolytic activity as well as proteolytic (caseinolytic) activity. The results of inhibitor studies and metal analyses strongly suggested that it was a zinc-protease. The purified HCE consisted of two probable isomers, HCE-1 and HCE-2. Both of them were markedly similar in amino acid composition, specific activities of choriolysis and proteolysis, and substrate specificity as determined using MCA-peptides. Moreover, they were not separable on SDS-PAGE, electrofocusing PAGE, or ultracentrifugal analysis, but were discriminated only on HPLC with a CM-300 column. Thus, the mixture of HCE-1 and HCE-2 could be regarded as almost a single enzyme, HCE. When it acted on an intact chorion, the purified HCE caused a remarkable swelling of its inner layer with concomitant release of peptides from it. Once the inner layer of chorion was swollen, the enzyme hardly digested it.     

Effects of low pH on the chorion of rainbow trout, Oncorhynchus mykiss, and brown trout, Salmo trutta f. fario

Morphological studies on the effect of low pH on egg shells (chorion) of rainbow trout, Oncorhynrhus mykiss Walbaum (formerly Salmo gairdneri ), and brown trout, Salmo trutta f. fario L., showed that the plugs of the outer layer of the chorion, which block the channels of the underlying layer, developed distinctive vacuolization at low pH exposure. This may lead to loss of chorional semi-permeability. Chorions were incompletely degraded at low pH inducing hatching problems. Partially hatched alevins showed only a small local zone of dissolution of the chorion around the head of the embryo preventing the larvae from being freed.     

Hatching mechanism and accelerated hatching of the eggs of a sac-spawning euphausiid Nematoscelis difficilis

Observation of the sac-spawning euphausiid Nematoscelis difficilisHansen during shipboard laboratory incubations showed that itsembryos usually hatch as pseudometanauplius (PMN) or metanauplius(MN). The eggs, which have a minute perivitelline space, arespherical at spawning and become elliptical after the nauplius1 develops. When ready to hatch, the PMN or MN embryos extendand contract their first and second antennae in a swimming movement,breaking the chorion into almost equal halves joined by onesmall section in the anterior part of the chorion. The mandiblesplay a secondary role in cutting the chorion. Then the embryopushes itself backwards with the first and second antennae toescape from the chorion. This is known as ‘push-off’hatching. The embryos always hatch progressively from the distalend towards the proximal end of the ovigerous sac. The timebetween hatching of the first and last embryo may reflect thetime the females require to lay a clutch of eggs (<2.1 h).Development time to the PMN stage at 10°C was     

Latent effects of egg incubation temperature on growth in the lizard Anolis carolinensis

Varied egg incubation temperatures can result in immediate effects on the phenotype of reptiles, and also latent effects that can augment or contradict effects evident at egg hatching. I examined the effects of incubation temperature on embryonic development, hatching morphology, and subsequent growth in multiple populations of the lizard Anolis carolinensis. Eggs from wild-caught females in four populations were incubated at up to three temperatures, 23.5, 27, and 30 degrees C. Measures of body size were collected immediately after hatching and weekly thereafter, while juveniles were maintained in a common laboratory environment for 8 weeks. Cooler incubation temperatures resulted in longer incubation periods but did not affect conversion of egg mass to hatchling mass. Incubation temperature did not affect hatchling mass or snout vent length (SVL), but did affect subsequent growth in both mass and SVL, which varied by population. Cooler incubation temperatures generally resulted in greater overall growth over 8 weeks of housing all juveniles in a common environment. In A. carolinensis, egg incubation temperature had latent effects on juvenile growth despite the absence of any detected immediate effects on hatchling phenotype. Therefore, the total impact and evolutionary importance of developmental environment should not be assessed or assumed based solely on the phenotype of reptiles at birth or hatching.     

Comparative analysis of the development of the lizard,Liolaemus tenuis tenuis. II. A series of normal postlaying stages in embryonic development

In this work, we have completed a study of the development of the ovoviviparous lizard Liolaemus tenuis tenuis. Ovoviviparity in this lizard is a condition in which eggs are retained within the reproductive duct for about 60 days. During this period the phases of segmentation, gastrulation, neurulation, presomitic, and somitic embryos transpire. During the months of December and January the eggs are laid, and at this time the embryos are comparable to stage 27 Liolaemus gravenhorsti lizard embryos, or to stage 29 Calotes versicolor lizard embryos. Differentiation of the facial region occurs between Days 12 and 42 after egg laying. Limbs develop rapidly between the 8th and 23rd days. By 53 days the appendicular skeleton is completely formed. After 36 days the mesonephros begins to degenerate, and its function is gradually taken over by the developing metanephros. Newborn lizards do not possess an egg caruncle. During the period up to hatching, there is a great increase of liquid within the egg, presumably amniotic fluid. Cracks develop in the leathery shell shortly before hatching and are, perhaps, the first sign of the onset of hatching. Increase of liquid in the egg during postlaying development accounts for its increase in weight and change in shape. Weight of the embryo at hatching does not exceed 32% of the total weight of the egg.     

Developmental consequences of association with a photosynthetic substrate for encapsulated embryos of an intertidal gastropod

Aggregation of embryos in clutches that lack internal circulation can increase the risk of hypoxia by limiting gas exchange. As a result, limits on oxygen solubility and diffusion in water can constrain the size and embryo concentration of aquatic egg clutches. Hypoxia in egg masses can slow embryo development, increase mortality, and reduce size at hatching. The risk of hypoxia for embryos, however, can be reduced by association with photosynthetic organisms. We examined whether embryo development in egg ribbons of the cephalaspidean mollusk Haminoea vesicula is significantly influenced by oviposition on eelgrass (Zostera marina). Association with the photosynthetic substrate had marked effects on development relative to association with non-photosynthetic substrates, and the direction of these effects was mediated by light conditions. Under intermediate and high light levels, association with eelgrass accelerated embryo development, while under dim light, the presence of the macrophyte increased development rate and reduced hatchling shell size. Benefits of association with eelgrass at higher light levels likely result from oxygen production by eelgrass photosynthesis, while we attribute costs under low light to oxygen depletion by eelgrass respiration. Association with Z. marina also limited microphyte growth in egg ribbons of H. vesicula. In the field, measurements of light attenuation within an eelgrass bed showed that conditions under which benefits accrue to embryos are ecologically relevant and correspond to spatial patterns of oviposition on eelgrass in the field. The choice of a photosynthetic oviposition substrate under appropriate light conditions can improve embryo fitness by accelerating embryo development without compromising hatchling size and by reducing the potential for excessive and harmful fouling by microphytes.     

Growth and uptake of mineral by embryos of the direct-developing frog Eleutherodactylus coqui

Embryos of the direct-developing frog Eleutherodactylus coqui take up small quantities of yolk and yolk mineral early in incubation but increase their uptake of yolk reserves at later stages of development. Growth and accumulation of calcium and magnesium by embryos also occur slowly at first and at a higher rate later. Accumulation of calcium and magnesium by embryos is largely a function of variation in size of embryos, but uptake of phosphorus is unrelated to size. Althrough patterns of growth and uptake of mineral by embryonic coquis resemble those for embryos of oviparous amniotes, embryonic coquis do not deplete the yolk of its nutrients to the same degree. Thus, residual yolk of coqui hatchlings contains a high percentage of the nutrient reserves originally present in the egg. This difference between embryonic coquis and embryos of oviparous amniotes may indicate that transfer of nutrients from yolk to embryo becomes limiting during the grwoth phase. Alternatively, some aspects of the neurologic system are so poorly developed at hatching that coqui may not be able to find prey effectively. A large nutrient reserve could sustain hatchling while the neurologic system continues to mature.     

The occurrence of the second moult of Ascaris lumbricoides and Ascaris suum

Maung M. 1978. The occurrence of the second moult of Ascaris lumbricoides and Ascaris suum. International Journal for Parasitology 8: 371–378. Eggs of Ascaris lumbricoides and A. suum were cultured at 28°C and observed daily. Larvae were released by pressure, by artificial hatching with CO₂, and by natural hatching after infection of laboratory mice. The early stages of development in the egg were observed to comprise two moults, one occurring immediately after the other. Both moults were initiated within the egg, but the time of completion of the second moult varied considerably, and in some instances was not completed until the larvae reached the liver of experimentally infected animals.     

人工养殖条件下嘉庚蛸的生物学特性及繁育技术初探

对人工养殖条件下嘉庚蛸(Octopustankahkeei)的基础生物学和繁育技术进行了初步研究。嘉庚蛸喜栖息于阴暗的场所,属昼伏夜出型,主要捕食一些活体的天津厚蟹(Helicetientsinensis)等。其个体产卵量非常低,为60～1 1 0粒头,平均仅85粒 头。繁殖季节,雌雄个体间存在交配行为,交配后精子贮藏在雌性的纳精囊中。受精卵分2～3批产出,成葡萄状悬挂于瓦罐底部,亲蛸(♀)具有护卵行为。受精卵约经过40～45d孵化出膜,为盘状卵裂,经过肉眼可见的“红珠”期、“黑珠”期、胚胎的逆转,最后孵化出幼体。     

Genetic variation in pathogen-induced early hatching of toad embryos

Accelerated hatching is one of few defences available to embryos, and is effective against many egg-stage risks. We present the first analysis of genetic variation in hatching plasticity, examining premature hatching of American toad embryos in response to pathogenic water moulds. We reared eggs from half- and full-sib families in the presence and absence of water mould. Hatching age and hatchling size showed low cross-environment genetic correlations, suggesting that early-induced hatching can evolve largely independently of spontaneous hatching. We found less phenotypic and additive genetic variation for early-induced hatching than spontaneous hatching, and a stronger correlation between egg and induced hatchling sizes. Directional selection by the pathogen may have eroded variation in early-induced hatching, pushing it against the constraint of hatching gland development. Later hatching has a second, muscular component. This pattern of variation may characterize defences based on developmental transitions, although other inducible defences show more variation in induced phenotypes.