Development and swimming behavior of Medaka fry in a spaceflight aboard the Space Shuttle Columbia (STS-107)

A space experiment aimed at closely observing the development and swimming activity of medaka fry under microgravity was carried out as a part of the S*T*A*R*S Program, a space shuttle mission, in STS-107 in January 2003. Four eggs laid on earth in an artificially controlled environment were put in a container with a functionally closed ecological system and launched on the Space Shuttle Columbia. Each egg was held in place by a strip of Velcro in the container to be individually monitored by close-up CCD cameras. In the control experiment, four eggs prepared using the same experimental set-up remained on the ground. There was no appreciable difference in the time course of development between space- and ground-based embryos. In the ground experiment, embryos were observed to rotate in place enclosed with the egg membrane, whereas those in the flight unit did not rotate. One of the four eggs hatched on the 8th day after being launched into space. All four eggs hatched in the ground unit. The fry hatched in space was mostly motionless, but with occasional control of its posture with respect to references in the experimental chamber. The fry hatched on ground were observed to move actively, controlling their posture with respect to the gravity vector. These findings suggest that the absence of gravity affects the initiation process of motility of embryos and hatched fry.     

Ten years after medaka fish mated and laid eggs in space and further preparation for the life-cycle experiment on ISS.

In the second International Microgravity Laboratory (IML-2/STS-65) mission in 1994, medaka fish (Oryzias latipes) performed their successful mating behavior in space for the first time among vertebrate animals. The eggs the fish laid in space developed normally, and hatched as fry (baby fish) in space. Those fry born in space had the same number of germ cells as the ground control fish, and these germ cells later developed to produce the offspring on the ground. Fry hatched in space did not exhibit any looping behavior regardless of their strain, visual acuity, etc. For fish, one of the most exciting experiments to be done abroad the International Space Station (ISS) is a realization of fish life cycles in microgravity. At present, fish are the most likely candidates to be the first vertebrate to live their life cycle in space. Research done in our laboratory for realizing the life-cycle experiment of medaka fish are also introduced.     

Induction of egg hatching by high humidity in the cicada Cryptotympana facialis

The timing of egg hatching in Cryptotympana facialis was examined in relation to short-term weather conditions. The brief underwater submergence, once a week, of dead twigs bearing cicada egg nests resulted in high hatching rates both at 25 degrees C and under outdoor conditions protected from rainfall. Under outdoor conditions with natural rainfall, most eggs hatched on rainy days. There was a significant positive correlation between the number of hatching eggs and daily relative humidity (RH). When eggs picked from twigs were exposed to various humidity levels, many eggs hatched quickly at higher humidity without direct contact with liquid water. Newly hatched nymphs showed a low tolerance to desiccation; at 81% RH at 25 degrees C, most of them died within 6 h. Under outdoor conditions, most nymphs died within 2 h on sunny days, whereas nymphs survived longer on rainy days. When newly hatched nymphs were released on dry ground, only 24% of them succeeded in burrowing into the soil, and many were killed by ants or desiccation. However, 92% of nymphs released onto wet ground successfully burrowed into it. The direct induction of hatching by high humidity ensures the survival and establishment in the soil of newly hatched nymphs in this species.     

The effect of exposure to microgravity on the development and structural organisation of plant protoplasts flown on Biokosmos 9

Preparatory experiments for the IML-1 (International Microgravity Laboratory) mission to be flown on the Space Shuttle in January, 1992, were performed on a 14 day flight on Biokosmos 9 (Kosmos 2044) in September 1989. The purpose of the experiment was to study the effect of weightlessness on protoplast regeneration. Problems with late access to the space vehicle meant that the newly isolated protoplasts from hypocotyl cells of rapeseed (Brassica napus L. cv Niklas) and suspension cultures of carrot (Daucus carota L, cv Nobo) had to be stored at 4 degrees C for 36 h prior to the launch of the biosatellite, in order to delay cell wall regeneration until the samples were in orbit. In the flight samples and the ground controls, a portion of the total number of protoplasts regenerated cell walls. The growth of flight rapeseed cells was only 56% compared to the ground control; the respective growth of carrot cells in orbit was 82% of the ground control. Analysis demonstrated that the peroxidase activity and the amount of protein was lower in the flight samples than in the ground controls. The number of different isoenzymes was also decreased in the flight samples. A 54% decrease in the production of cellulose was found in rapeseed, and a 71% decrease in carrot. Hemicellulose production was also decreased in the flight samples compared to the ground controls. Ultrastructural analysis of the cell aggregates from the protoplasts cultured in orbit, demonstrated that hydrolysis and disappearance of reserve starch occurred in the flight cell plastids. The mitochondria were more varied in appearance in the flight samples than in the ground control cells. An increased frequency of the occurrence of folds formed by the plasmalemma together with an increase in the degree of complexity of these folds was also observed. Fluorescence analysis showed a decrease of the calcium content in cell cultures under space flight compared to the ground controls. One general effect of the stay onboard the space vehicle was a retardation of the regeneration processes. Callus cultures obtained from the flight samples grew very slowly compared to callus regenerated from the ground controls, and two years after the Biokosmos 9 flight there appears to be no further growth in the samples exposed to microgravity. Callus cultures from the ground controls, however, continue to grow well. A simulation experiment for IML-l performed in January 1990 at ESTEC (European Space Technology Center), The Netherlands, has resulted in regenerated plants. These observations are discussed and compared to the results obtained on Biokosmos 9.     

Mortality factors of eggs of Leptocorisa chinensis (Hemiptera: Alydidae) in rice fields

To clarify factors causing mortality of Leptocorisa chinensis Dallas (Hemiptera: Alydidae) eggs in rice fields, sentinel egg masses were exposed for seven days in two rice, Oryza sativa L. (Poaceae), fields. An insecticide was sprayed to remove natural enemies in one-half of each field before exposing egg masses to predation. An egg mass containing 14 eggs was glued to a plastic label, and 21 egg masses each were placed in the sprayed and unsprayed control plots. During exposure, the number of hatched and missing eggs was observed daily. Egg predators were sampled in the fields before and after insecticide application. After the egg masses were retrieved from the fields, mortality factors of the unhatched eggs were assessed in the laboratory. The mean number of hatched and missing eggs was not significantly different between the sprayed and control plots in field A. In field B, however, the numbers were significantly different. The percentage of missing eggs in damaged egg masses ranged from 80 to 100%. In the laboratory, we observed that feeding marks caused by the grasshopper Conocephalus chinensis (Redtenbacher) were similar to those on the eggs exposed in the rice fields. The density of C. chinensis was low in control plots of field A. In contrast, the density was high in those of field B. These observations suggest that the density of egg predators, e.g., C. chinensis, is a mortality factor of L. chinensis eggs in rice fields.     

Behavior and reproduction of invertebrate animals during and after a long-term microgravity: space experiments using an Autonomous Biological System (ABS).

Aquatic invertebrate animals such as Amphipods, Gastropods (pond snails), Ostracods and Daphnia (water flea) were placed in water-filled cylindrical vessels together with water plant (hornwort). The vessels were sealed completely and illuminated with a fluorescent lamp to activate the photosynthesis of the plant for providing oxygen within the vessels. Such ecosystem vessels, specially termed as Autonomous Biological System or ABS units, were exposed to microgravity conditions, and the behavior of the animals and their reproduction capacity were studied. Three space experiments were carried out. The first experiment used a Space shuttle only and it was a 10-day flight. The other two space experiments were carried out in the Space station Mir (Shuttle/Mir mission), and the flight units had been kept in microgravity for 4 months. Daphnia produced their offspring during a 10-day Shuttle flight. In the first Mir experiment, no Daphnia were detected when recovered to the ground. However, they were alive in the second Mir experiment. Daphnia were the most fragile species among the invertebrate animals employed in the present experiments. All the animals, i.e., Amphipods, pond snails, Ostracods and Daphnia had survived for 4 months in space, i.e., they had produced their offspring or repeated their life-cycles under microgravity. For the two Mir experiments, in both the flight and ground control ecosystem units, an inverse relationship was noted between the number of Amphipods and pond snails in each unit. Amphipods at 10 hours after the recovery to the ground frequently exhibited a movement of dropping straight-downward to the bottom of the units. Several Amphipods had their legs bent abnormally, which probably resulted from some physiological alterations during their embryonic development under microgravity. From the analysis of the video tape recorded in space, for Ostracods and Daphnia, a half of their population were looping under microgravity. Such looping animals could be observed still at the end of the 4 month stay in space. No looping behavior was noted for Amphipods and pond snails.     

Performance of a spring-feeding moth in relation to time of oviposition and bud-burst phenology of different host species

Abstract. 1. As a spring-feeding moth committed to immature foliage, the autumnal moth Epirrita autumnata (Lepidoptera, Geometridae) must have egg hatch synchronised with the bud-burst of its host plants. Due to large individual variation in the length of the pupal period, however, E. autumnata populations exhibit a prolonged period of flight and oviposition. Because the timing of oviposition in autumn is associated with the timing of egg hatch in the following spring, the time window for egg hatch expands and more potential hosts may become attainable. This suggestion was evaluated under field conditions by rearing E. autumnata eggs and larvae on four different hosts.
2. The performance of E. autumnata was measured by using estimates for fecundity (pupal mass) as well as survivorship of eggs and larvae. Based on the availability of foliage and phenological synchrony between larval and leaf development, early-laid eggs and the larvae originating from them were predicted to perform better on the hosts that have early-flushing leaves. On the late-flushing hosts, the larvae that hatched later were predicted to perform better than the larvae that hatched earlier. Half of the trials were exposed to predators and parasitoids, while the rest were conducted inside mesh-bags preventing larval dispersal and mortality due to natural enemies.
3. The results of the experiment did not support the simple predictions. In particular, host-plant quality and natural enemies appeared to operate discordantly between early- and late-laid eggs. Larvae from the late-laid eggs had rapid development during the larval stages and pupated at the same time and with the same pupal mass as the larvae hatched from the early-laid eggs.
4. The results indicate an occurrence of several, unknown selective forces in E. autumnata populations maintaining variation in the length of the pupal period, timing of oviposition, and timing of egg hatch.     

Trichogramma releases in North Carolina cotton: why releases fail to suppress heliothine pests

Field studies were conducted in 1996 and 1997 to determine the fate of naturally oviposited F3 heliothine eggs in cotton plots treated with augmentative releases of Trichogramma exiguum Pinto & Platner and nontreated plots. Four cohorts of newly oviposited eggs (< 24 h old) were followed in 1996 and two cohorts in 1997. In 1996, mean +/- SD percent parasitism, estimated by in-field studies following the fate of naturally oviposited eggs, ranged from 7 +/- 7 to 61 +/- 8% in T. exiguum release plots and 0 +/- 0 to 35 +/- 13% in control plots. The mean +/- SD percent of eggs hatched in T. exiguum release plots ranged from 1 +/- 2 to 11 +/- 4% and 7 +/- 4 to 28 +/- 10% in control plots. In 1997, mean +/- SD percent egg parasitism ranged from 27 +/- 4 to 40 +/- 3% in T. exiguum release plots and 15 +/- 18 to 25 +/- 8% in control plots. The mean +/- SD percent of eggs hatched in T. exiguum release plots ranged from 7 +/- 3 to 12 +/- 2% and 18 +/- 6 to 28 +/- 8% in control plots. Despite increased parasitism and reduced egg hatch in T. exiguum release plots, overall, there was no significant difference in larval density (all instars combined) between T. exiguum release and control plots. Combined analysis of the heliothine larval populations and egg fate data revealed that the additional egg mortality produced by released T. exiguum was offset by lower larval mortality in release plots. Because of the occurrence of compensatory mortality, the egg stage of heliothines is not an appropriate target for biological control using Trichogramma wasp releases.     

Egg size and offspring fitness in a bdelloid rotifer

To test if the quality of offspring is affected by egg size and whether it increases with the amount of resources allocated to an individual egg, we compared the offspring hatched from small and large eggs of Macrotrachela quadricornifera(Rotifera, Bdelloidea). Differently sized eggs were obtained by feeding mothers with different food concentrations. Large eggs were expected to provide better offspring in terms of (1) development time of the embryos, (2) age at first reproduction of the juveniles and (3) recovery after desiccation of the newborns. The comparison between offspring hatched from large vs. small eggs of the same bdelloid species revealed that animals hatched from large eggs had shorter embryonic development and earlier age at first reproduction than animals hatched from small eggs. In contrast, the capacity to survive stress, like desiccation, was not affected by egg size. Thus, offspring hatched from larger eggs had shorter generation times, but were not favoured under harsh circumstances.     

Egg hatching of two locusts,Schistocerca gregaria and Locusta migratoria,in response to light and temperature cycles

The present study showed that the eggs of the desert locust, Schistocerca gregaria, and the migratory locust, Locusta migratoria, responded to photoperiod by hatching when placed on sand in the laboratory. S. gregaria mainly hatched during the dark phase and L. migratoria during the light phase. The importance of light as a hatching cue depended on the magnitude of the temperature change during the thermoperiod; photoperiod played a more important role in the control of hatching time in both species when the magnitude of the temperature change was small. In addition, the eggs of the two species that were covered with sand did not respond to photoperiod and hatched during both the light and dark phases, indicating that light did not penetrate through the sand. Because locust eggs are normally laid as egg pods and a foam plug is deposited between the egg mass and the ground surface, we tested a possibility that naturally deposited eggs perceived light through the foam plug. The eggs that were deposited and left undisturbed in the sand hatched during the light and dark phases at similar frequencies. These results suggest that the eggs of both locust species responded to light and controlled their hatching timing accordingly but would not use light as a hatching cue in the field. The evolutionary significance of the ability of eggs to respond to light in these locusts was discussed.     

STS-95 Space Experiments (plants and cell biology).

We report the outline of Space Experiments conducted on Space Shuttle (STS-95) launched in autumn of 1998. In this STS-95 mission, Japanese astronaut Dr. Chiaki Mukai achieved her 2nd space flight and conducted a part of 82 space experiments including Japanese experiments. US astronaut Senator John Glenn also achieved his second space flight, 36 years after his first space flight. Senator Glenn was a leader of the original (the first) 7 US astronauts and very famous in US because he succeeded US first orbital space flight around the earth. NASDA had started the project of space experiment using STS-95 at the summer of 1997, therefore we had only one year for the all preparation Yamashita, et al. Biological Sciences in Space, Vol.12 No.3(1998). Scientific results will be reported by investigators, therefore we report here how we had been developing the space experiment plan, on board operation procedure and ground operations including ground control experiments about four plant experiments and one cell biology experiment.     

颗粒体病毒在小菜蛾种群中的垂直传播方式的初步观察

在室内观察了小菜蛾颗粒体病毒PlutellaxylostellaGranulosisVirus(RxGV)在小菜蛾种群水平上可能的垂直传播途径。结果表明 :(1 )上代小菜蛾Plutellaxylostella(L .)幼虫期饲毒不能将小菜蛾颗粒体病垂直传递到下代 ;(2 )取食病毒的雌蛾产的卵所孵化的幼虫中 ,没病毒感染出现 ,说明病毒不能经小菜蛾成虫体内传播到下一代。 (3 )用小菜蛾颗粒体病毒处理卵卡时 ,其初孵幼虫有少量被病毒感染死亡 ,浓度高时感染率也较高 ,但感染率均在 1 0 %以下 ,说明小菜蛾颗粒体病毒能经卵表垂直传递给下代 ,但传递效率不高。但当用病毒处理叶片上的卵时 ,初孵幼虫死亡率高达 77 73 % ,而且孵化出的幼虫大多数染病且死于 3龄以前 ,因此田间病毒的施用时间可提早到卵高峰期。     

Effect of weight and storage time of broiler breeders’ eggs on morphology and biochemical features of eggs,embryogenesis, hatchability,and chick quality

The transfer of hatchability results obtained under experimental conditions to the commercial ground with a positive financial effect proves the value and usefulness of these data. On the other hand, finding results on commercial processes of broiler breeders’ egg incubation in the literature is challenging. The presented study aimed to determine the effects of egg weight and storage time on the physical, biochemical characteristics of hatching eggs, embryogenesis and hatchability in Ross 308 broiler breeders. On the laying day, the eggs were divided into four weight groups: S – small eggs (57–61 g), M – medium eggs (62–66 g), L – large eggs (67–71 g), and XL – extra-large eggs (72–76 g). The eggs were then stored for 3, 7, 14, and 21 days under controlled conditions. As the egg storage time increased, a decrease in the yolk quality (lower index) was observed. The highest Haugh units were found in eggs from the S and M groups. The cholesterol content of the M, L, and XL groups was lower on days 7, 14, and 21 as compared to that of eggs only stored for 3 days. Egg weight loss during incubation decreased with an increase in the egg weight. An extension of the egg storage time caused an increase in the loss of egg weight. On the 14th and 18th days of hatching, an increase in the eggshell temperature was noted with an increase in the weight of the egg. The eggs stored for 7 days were characterised by the highest shell temperature on each day. The highest hatchability percentage was recorded for the M group. The hatchability rate decreased with the prolongation of the storage time, while the number of crippled chicks after hatching increased. The results confirmed that the increased weight of the eggs and prolonged storage time (14 and 21 days) increased the weight and decreased the length of the newly hatched chicks, respectively. Chicks from the heaviest eggs and those stored for 14 and 21 days showed poor results on the Pasgar score® test. The observations indicate the need to adopt various (of those available) methods to assess the quality of newly hatched chicks in hatcheries in order to produce high-quality broiler chickens. The results also indicate that prolonged egg storing beyond 14 days may affect the thyroid hormone economy during the hatching of chicks, especially in the XL group.     

Hatching and viability of rotifer diapausing eggs collected from pond sediments

1. Planktonic rotifers inhabiting variable environments produce diapausing eggs that accumulate in the sediment of lakes and ponds, forming egg banks that may withstand adverse periods. A common assumption in zooplankton diapausing egg bank studies is to count as viable all eggs in the sediment that look healthy. This assumption should be challenged by asking how effectively ‘healthy‐looking’ eggs represent viable eggs. 2. In this study, viability of more than 1100 ‘healthy‐looking’ diapausing eggs belonging to the Brachionus plicatilis species complex was assessed in a laboratory hatching experiment. Eggs were collected at different depths from sediment cores obtained from 15 ponds located in coastal and inland areas of Eastern Spain. 3. Only approximately one half of the ‘healthy‐looking’ diapausing eggs hatched after incubation in experimental conditions. Almost all the hatchlings (99.4%) survived to maturity. The proportion of ‘healthy‐looking’ diapausing eggs that hatched varied among areas and among ponds within area, and substantially declined with sediment depth. Most of the hatchlings (88%) were obtained from the uppermost 2 cm of sediment. ‘Healthy‐looking’ eggs from upper sediment layers hatched after significantly shorter incubation times than eggs recovered from deeper layers. 4. Both decreased hatching success and increased incubation time for hatching with sediment depth suggest that older ‘healthy‐looking’ eggs are less responsive to hatching stimuli and could become unviable. However, the strong correlation found between the number of ‘healthy‐looking’ eggs and the number of hatchlings indicates that the abundance of ‘healthy‐looking’ eggs is a good index of egg bank viability.     

Characterization and functions of the whitefly egg pedicel.

For the silverleaf whitefly, Bemisia argentifolii (Bellows and Perring) (Homoptera: Aleyrodidae), scanning and transmission electron microscopic techniques were used to observe the characteristics of egg oviposition into both plant cells/tissues and artificial membranes, and to document the morphology of mature egg pedicles removed from the ovaries of females. The exterior of the distal portion of the pedicel consisted of a tangled array of fibrous structures (0.2-0.3 microm in diameter) that constituted about 20-25% of the outer diameter of the pedicel. The attachments of the fibers to the core of the pedicle suggested that the pedicel functions as the collector and conduit for water (vapor), and perhaps solute movement into the egg. Silverleaf whitefly eggs on membranes were incubated at various levels of relative humidity and the eggs were scored for egg hatch. At 98-100% rh, the percentage egg hatch was 86-98%. At lower humidity ranges of 0-20, 55-65, and 75-85% rh, none of the eggs hatched. Media (solute) uptake by silverleaf whitefly egg pedicels was determined by exposing the pedicel side of eggs oviposited on membranes to media solutions containing the high molecular weight polysaccharide, [(14)C]-inulin. Solute uptake by the pedicel and movement into developing silverleaf whitefly eggs were demonstrated using [2-(14)C]-acetate, and assaying for radioactivity in hatched nymphs. These studies, using exposure of pedicels to relative humidity and radiolabeled materials, demonstrate that whitefly egg hatch is dependent upon water uptake by the pedicel, and that the pedicel has the ability to transport solutes into the developing egg.     

Corticosterone stimulates hatching of late-term tree lizard embryos

The regulation of hatching in oviparous animals is important for successful reproduction and survival, but is poorly understood. We unexpectedly found that RU-486, a progesterone and glucocorticoid antagonist, interferes with hatching of viable tree lizard (Urosaurus ornatus) embryos in a dose-dependent manner and hypothesized that embryonic glucocorticoids regulate hatching. To test this hypothesis, we treated eggs with corticosterone (CORT) or vehicle on Day 30 (85%) of incubation, left other eggs untreated, and observed relative hatch order and hatch time. In one study, the CORT egg hatched first in 9 of 11 clutches. In a second study, the CORT egg hatched first in 9 of 12 clutches, before vehicle-treated eggs in 10 of 12 clutches, and before untreated eggs in 7 of 9 clutches. On average, CORT eggs hatched 18.2 h before vehicle-treated eggs and 11.6 h before untreated eggs. Thus, CORT accelerates hatching of near-term embryos and RU-486 appears to block this effect. CORT may mobilize energy substrates that fuel hatching and/or accelerate lung development, and may provide a mechanism by which stressed embryos escape environmental stressors.     

Capillaria hepatica infection of Mastomys natalensis: on the development, egg production and host reaction (author's transl)]

Studies on the development and egg production of Capillaria hepatica and on the macroscopically visible alterations of the liver and spleen of the host were carried out in experimentally infected Mastomys natalensis. Following the oral administration of infective eggs the first stage larvae hatched in the caecum of fasting and fed animals after about 8 and 15 hours respectively. The prepatency could be found with 20 days. The dynamics and duration of egg production of the parasite proved to be dependent on the infective dose. After the first week of patency increasing numbers of eggs/liver were found with increasing doses of infection up to 800 eggs per animal. This relation could not be observed 76 days post infection. After infections with 50, 300 und 800 eggs per animal maximum egg production was found between 60 and 72, 36 and 48 and about 30 days p.i., respectively. The egg production of the parasites correspondently continued for more than 85 days after infection or had ceased 72 or 48 days p.i. Intraperitoneal administration of infective eggs revealed a lower infection rate, evaluated by the number of eggs per liver, than oral infection.     

Environmental factors associated with the distribution of floodwater mosquito eggs in irrigated fields in Wrocław,Poland

A survey of distribution patterns of floodwater mosquito eggs related to environmental conditions such as moisture and plant associations was conducted by using soil samples from irrigated fields in Wroc?aw, Poland. Mosquito egg distribution was determined by repeatedly flooding the soil samples with aerated water at a temperature of 25° C. Under laboratory conditions, hatching in installments of Aedes caspius (Pallas) and Aedes vexans (Meigen) were commonly observed. The results show that ～75% of the larvae of Ae. caspius and Ae. vexans hatched after the first flooding under summer‐like conditions, whereas, following the second and third flooding, the numbers of hatched larvae were significantly lower. In our study, within one intermediate flooded field, a total of 66 plant species was identified and classified into six communities. All vegetation types were associated by varied egg densities and showed differences both in richness and Shannon‐Wiener diversity index as well as in ecological indices for moisture, soil reaction, and nutrient level. Small changes in elevation along the slope within the study area showed a large difference in the distribution of mosquito eggs. The highest average egg density was observed in zones with high occurrence of Phalaris arundinacea, usually prevalent in intermediate flooded and fertile areas. Knowledge of the indicators for the distribution of floodwater mosquito eggs in temporary breeding sites may be essential for organizing a successful, integrated mosquito control program with special regard to microbial control agents.     

Life Cycle and Mating Behavior of Belonolaimus longicaudatus in Gnotobiotic Culture

The life cycle of Belonolaimus longicaudatus was observed in vitro on excised roots of Zea mays. Roots were cultured on Gamborg''s B5 medium in petri dishes with 1.5% agar adjusted to pH 5.8 and incubated at 28 °C in darkness. Second-stage juveniles (J2) fed on the roots and started the second molt (M2) to the third-stage juveniles 2 days after inoculation (DAI). The third molt (M3) to the fourth-stage juveniles occurred 7 DAI, followed by the fourth molt (M4) to males 13 DAI or to females 14 DAI. Nematode gender differences were observed by the end of the fourth molt. The first male appeared 15 DAI and the first female 17 DAI, after which mating occurred. Males were attracted to females, and mating was observed. Mating was required for reproduction. Fertilized females began to lay eggs 19 DAI and continued egg laying without the further presence of males during a 90-day observation. All of the eggs hatched. Unfertilized females rarely laid eggs, and none of the eggs were able to hatch. Feeding took place between each molt and before egg deposition occurred. The first-stage juveniles molted in the eggs 4 days after deposition, and J2 hatched from eggs 5 days after egg deposition. The life cycle from J2 to J2 was completed in 24 days.     

Planktonic egg masses of the diamond-shaped squid Thysanoteuthis rhombus in the eastern Atlantic and the Mediterranean Sea

Eight planktonic egg masses of the diamond-shaped Thysanoteuthisrhombus observed from 1995 to 2000 are described. Four werefound in the western Mediterranean and the others were foundoff the Canary Islands. The egg masses from the Canary Islandsare the first records for the eastern Atlantic. All were foundnear the surface at the end of summer and the beginning of autumn.The planktonic egg masses were dense, resilient oblong cylinderswith rounded tips ranging from 80 to 130 cm in length and between15 and 20 cm in diameter. Egg capsule dimensions ranged from2.8 to 3.4 mm and total length of the newly hatched paralarvaewas between 2.5 and 2.8 mm. Each egg mass contained an estimated24 100–43 800 eggs. Some new characters that should helpidentification of the paralarvae, such as arm formulae, presenceof an incipient keel-shaped membrane on some arms, and the typeand chromatophore pattern are given. Ecological factors influencingthe presence and distribution of these egg masses are discussed.