Cloning and expression of ecdysone receptor (EcR) from the intertidal copepod, Tigriopus japonicus

Ecdysteroids are steroid hormones that play an important role in development, growth, molting of larva, and reproduction in the Arthropoda. The effect of ecdysteroids is mediated by its binding to ecdysteroid receptor (EcR). To investigate the role of EcR during development and the effect to environmental stressors on EcR expression in a copepod, we isolated and characterized cDNA and 5′-promoter region of the Tigriopus japonicus EcR (TJ-EcR), and studied mRNA expression pattern. The full-length TJ-EcR cDNA sequence was 1962 bp in length and the open reading frame encoded 546 amino acids. The deduced TJ-EcR protein contained well-conserved DNA-binding domain and ligand-binding domain. Phylogenetic analysis revealed that TJ-EcR was clustered with the EcR of other crustaceans. TJ-EcR mRNA was expressed in a developmental stage-specific manner: high in early developmental stages and low in the adult stage. Significantly elevated expression of the TJ-EcR gene in adults was detected at hypersalinity (42 ppt) and high temperature (35 °C) condition. The 5′-flanking region of TJ-EcR gene contains heat shock protein 70 response elements, implying that the environmental stressors may affect its expression via the stress-sensor. In addition, bisphenol A (100 µg/L) repressed TJ-EcR expression. Our results suggest that TJ-EcR could be a biomarker for the monitoring of the impact of environmental stressors in copepods.     

The effect of food limitation on gonad development and egg production of the planktonic copepod Calanus finmarchicus

Calanus finmarchicus, one of the dominant copepods species of the North Atlantic, often encounters low food concentration or quality during the reproductive period; however, our knowledge on the effect of these conditions on reproduction processes is scarce. The present study combines experiments with histological observations to describe the response of C. finmarchicus to limited food focussing on (1) oocyte maturation processes, (2) gonad morphology, (3) egg production rates as a function of spawning frequency and clutch size, and (4) the fuelling of egg production by measuring carbon and nitrogen content of the females. In the laboratory, C. finmarchicus females were exposed to 0, 10, 50, 150 or >300 μg C l⁻¹ for several days. To account for food quality and season, reproductive activity was compared in April and July 1999 between females feeding on diatoms or dinoflagellates. The effect of feeding history was studied in February with females fed and starving prior to the experiment. Feeding conditions had severe effects on oocyte maturation process. Hence, egg production varied significantly with food concentration and quality, season and feeding history due to variation in both clutch size and spawning frequency. Clutch size differed by a factor of 2-4 between food limited and well fed females, and is thus an important parameter for modelling egg production. Changes in clutch size were related to changes of the number of maturing oocytes in the females gonads indicating that the latter can be used to precise the prediction of egg production from preserved samples. The proportion of females carrying at least some mature oocytes was relatively high at low food availability. Apparently, these females used internal body reserves as the carbon and nitrogen content decreased significantly under these conditions. These results indicate that C. finmarchicus embarks on the strategy to enable reproduction in all or many females of a population at low rates when feeding conditions are unfavourable.     

Effects of copulation temperature on female reproductive output and longevity in the wolf spider Pardosa astrigera (Araneae: Lycosidae)

The copulation duration of male wolf spider Pardosa astrigera, was significantly influenced by environmental temperature, as had been found in some insect species. Therefore, temperature during male courtship and copulation may influence the amount of sperm and seminal fluids transferred during copulation, which in turn could influence female fitness. In order to test this hypothesis, we subjected pairs of male and female P. astrigera to five temperature groups from 16 to 32 °C at an interval of 4 °C, and investigated whether and to what extent the various temperatures during male courtship and copulation influenced female reproductive output and female adult longevity under controlled laboratory conditions. With the increase of copulation temperature, females were more likely to lay egg sacs. The total egg sacs and lifetime fecundity of female were positively influenced by copulation temperature, whereas female lifetime spiderlings and adult longevity were independent of copulation temperature.     

Short-term impacts of polyunsaturated aldehyde-producing diatoms on the harpacticoid copepod, Tisbe holothuriae

Diatoms that produce toxic oxylipins can be detrimental to the reproductive success of aquatic invertebrates. Despite the potential importance of these toxins in shaping aquatic ecosystems, marine studies to date have focused almost exclusively on planktonic calanoid copepods. The current work examines the response of the benthic harpacticoid copepod, Tisbe holothuriae, to direct exposure to diatom-derived oxylipins and the short-term impact of oxylipin-producing diatom diets on reproductive success. The most toxic oxylipin was the polyunsaturated aldehyde (PUA) 2E,4E-decadienal with an LD₅₀ of 9.3 μM for T. holothuriae nauplii. The least tolerant life-stage was the nauplius followed by adult males then adult females. Short-term exposure to PUA-producing diatoms (Skeletonema marinoi and Melosira nummuloides) in maternal diets had no significant impact on reproductive success compared with non-PUA-producing diets (Skeletonema costatum, Navicula hanseni, Phaeodactylum tricornutum and Tetraselmis suecica). The PUA producers had no negative impact on the survival and development of naupliar stages to adulthood. T. holothuriae expresses a higher degree of tolerance to PUA-producing diatoms than many planktonic calanoids, possibly reflecting a degree of adaptation to higher stress levels associated with the benthos. This is the first study to investigate the reproductive responses of harpacticoid copepods feeding on known PUA-producing diatoms.     

The reproductive output of sea lice Caligus rogercresseyi under controlled conditions

Gravid females of Caligus rogercresseyi were collected from Atlantic salmon (Salmo salar) from a farm located at Chiloe Island (42°40′S73°15′W), Chile, to obtain information about the reproductive output of this parasite in vitro. The egg strings removed from the females were incubated under controlled conditions to obtain virgin adult females. One female which had mated only once produced eleven generations of eggs strings in a period of 74 days. The first egg strings of the females obtained in vitro were produced at 389 degree days (°D) after egg incubation, while the next generations of eggs strings were produced with a periodicity between 4 and 6 days dependent on the water temperature. The average length of the egg string was 3.1 mm and the mean number of eggs per string was 31. The values recorded in captivity for the egg string length and the number of eggs per string, were lower than the values recorded in gravid females from the field. One female survived for 79 days and males, maintained separately from the females, survived for 60 days.     

Effects of various environmental conditions on growth and reproduction of the sea hareAplysia oculifera (Adams and Reeve, 1850)

We studied the influences of food type, food quantity, water currents, starvation and light on growth and reproduction of the sea hareaplysia oculifera (Adams and Reeve, 1850) under laboratory conditions. Out of five species of algae served as food,Enteromorpha intestinalis promoted the fastest growth ofA. oculifera, Ulva spp. slower growth,Cladophora sp. allowed maintenance spp. slower growth,Cladophora sp. allowed maintenance of steady body mass, and the brown algaeColpomenia sp. andPadina pavonia were rejected by the sea hares. When sea hares were exposed to four levels of water currents, growth rates decreased as water currents increased. Sea hares fed on 50% ration grew slower than those fed on 100% ration (ad libitum). During 10 days of starvation sea hares lost weight, but when subsequently fed 100% ration they recovered and grew at a rate similar to those fed continuously with 100% ration. Under shade and under natural sunlight sea hares grew at the same rates. Whenever growth rates decreased, sea hares began to spawn at a smaller body size.A. oculifera demonstrated physiological plasticity that adapted them to varied and unpredictable environmental conditions. At different conditions of food availability they applied different tactics of resource allocation between growth and reproduction.     

Fecal cortisol levels in free-ranging female chacma baboons: relationship to dominance, reproductive state and environmental factors

Savannah baboons are one of the few mammalian species that do not exhibit seasonal reproduction patterns and are therefore ideally suited to study the effect of female reproductive states (cycling, pregnant, lactating) on cortisol levels independent of seasonal factors. Fecal samples from 10 free-ranging female chacma baboons (Papio hamadryas ursinus), collected during a period of 17 months, were analyzed using a steroid-extraction method. Reproductive state had a significant effect on fecal cortisol, with lowest levels found in estrous females. Fertility was not related to fecal cortisol levels; we found no significant differences between samples collected on conceptive and nonconceptive cycles. Environmental factors explained most of the variance of fecal cortisol levels. Cortisol measures were strongly correlated with seasonal differences such as daylight duration, temperature and the amount of time that baboons spent resting. We measured higher cortisol levels during winter months and suggest that this could be related to shorter resting periods and to the cold minimum ambient temperatures at this study site. Finally, we found no relationship between social rank nor the rate of agonistic interactions with basal fecal cortisol levels.     

Effects of different diets on the reproduction and naupliar development of the copepod Acartia bifilosa

The influence of diatoms on the reproduction and naupliar development of Acartia bifilosa was investigated under laboratory conditions, comparing initial in situ values and laboratory-food treatments. Egg production by A. bifilosa was significantly reduced by one diatom diet (Phaeodactylum tricornutum: Pt) and by two non-diatom diets (Platymonas subordiformis: Ps and Nannochloropsis oculata: No). It was less affected by the other diatom diet (Skeletonema costatum: Sc) or by two mixed-food treatments (D-mix and DG-mix), composed of two diatoms (Pt, Sc) and four species (Pt, Sc, Ps and No), respectively. The negative effect of Pt was eliminated when adult copepods were offered mixed-food diets. There were no significant differences between the hatching success values observed in filtered seawater and in algal exudates, indicating that diatoms did not produce active dissolved toxic substances under the different food concentrations tested. The mortality rate of nauplii was higher with Pt than the other diets, suggesting that this diatom species had a negative effect on egg production, hatching success and naupliar survival simultaneously. Compared to other diets, No and Pt were not beneficial food sources for reproduction and for female and larval survival.     

The reproductive cycle of the asteroid Coscinasterias muricata in Port Phillip Bay, Victoria, Australia

The reproductive cycles of two populations of Coscinasterias muricata from Port Phillip Bay, Victoria, Australia are described in terms of organ indices, oocyte development and progesterone levels. Both Governor's Reef and South Channel Fort populations exhibited clearly defined reproductive cycles with two spawning periods, during summer and during spring. In both populations, the pyloric caecal index and gonadal index showed inverse cycles suggesting nutrient translocation from the pyloric caeca to the gonads for gametogenesis. Physiological changes, such as decreases in pyloric caecal index, and increases in gonadal index and oocyte diameter were observed following the months of increased progesterone concentrations in the female pyloric caeca. These results indicate a role for this steroid in the reproductive cycle. The results from the Governor's Reef population suggest that the influence of progesterone on the reproductive cycle of female C. muricata is regulated by photoperiod. Results also provide evidence of a role for seawater temperature in the reproductive process of this species.     

The effect of photoperiod on the reproductive development of the northern bay scallop, Argopecten irradians irradians

One-year-old bay scallops, Argopecten irradians irradians (58 ± 2 mm, 22 ± 1 g live weight) were exposed to four replicated photoperiod treatments (24D, 8L:16D, 16L:8D, and 24L where D = dark hours, L = light hours) in order to measure the effect on gonad weight and maturation during the conditioning process. Results indicated that day-lengths of more than 8 h are necessary to promote gonad maturation in bay scallops. After 6 wk, the mean gonad weight for scallops in the 16-h and 24-h light regimes was similar at 0.6 ± 0.1 g dry weight compared to a mean of 0.2 ± 0.1 g dry weight for those in the 8-h and 0-h light regimes. Histological assessment indicated significantly more follicular tissue development in both the male and female portion of the gonad in the two longer photoperiod treatments. Overall, gamete maturity was highest for the scallops in the 16-h light regime; the incidence of mature eggs was 50% compared to 35% in the 24-h light regime, 20% in the 8-h light regime and 10% in the 0-h light regime. Assessment of feeding rates indicated no significant difference in algal cell consumption among treatments. Total dry tissue weight doubled over the 6-wk conditioning trial with no significant differences among treatments. One-year-old bay scallops appear to be non-responsive to conditions suitable for gonad maturation (i.e. appropriate temperature and food levels) unless they receive more than 8 h of light exposure. This finding has important implications for northern hatcheries which typically condition broodstock indoors during the early spring.     

Schistosoma japonicum: Tsunagi/Y14 protein plays a critical role in the development of the reproductive organs and eggs

Tsunagi/Y14 is an evolutionarily conserved RNA-binding protein that is required for the maintenance of oogenesis and the masculinization of the germ-line in many animal models. We speculated that Tsunagi/Y14 might also regulate reproductive organ development in Schistosoma japonicum (S. japonicum, Sj). Sj Tsunagi/Y14 and control double-stranded RNAs were introduced into schistosomula by electroporation respectively. These transfected schistosomula were cultured in vitro for 1, 3 or 5 days. The mRNA and protein levels of the target gene in the cultured schistosomula were significantly suppressed compared with those of the control group. Furthermore, BALB/c mice were infected with the transfected schistosomula for 6 weeks and were sacrificed to harvest the adult worms. We found that the silencing of Sj Tsunagi/Y14 led to defects in reproductive organs development in both male and female worms. Moreover, it also affected the size, quantity and activity of the eggs in the mice liver. Our findings indicated that Tsunagi/Y14 plays a critical role in the development of reproductive organs and eggs in S. japonicum.     

The reproductive response of the protobranch bivalve Yoldia hyperborea to an intermittent influx of phytodetritus. An experimental approach

Yoldia hyperborea (Loven) is a deposit feeder species living in muddy sediments of Conception Bay (Newfoundland, Canada) where it is exposed to a seasonal input of sinking phytoplankton during spring. Data field indicates that this species exhibits an increasing gonad development shortly after the sinking of phytodetritus event, suggesting a close relationship between this food pulse and reproduction. Laboratory experiments carried out with a mixture of laboratory-grown senescent algae Isochrysis galbana, Tetraselmis suecica and Chaetoceros affinis were able to mimic the reproductive response described for Yoldia individuals inhabiting Conception Bay.A dual marker experiment using ¹⁴C and ⁶⁸Ge radiolabelled Thalassiosira nordenskioldii demonstrated that Y. hyperborea fed on T. nordenskioldii and 14C was incorporated into the gonad tissue. Consequently experimental individuals periodically supplied with senescent algae concentrate produced more and larger oocytes than control individuals. These results would help to understand the influence of the sinking phytodetritus in the reproductive activity of Y. hyperborea at Conception Bay.     

Trophodynamics of two interacting species of estuarine mysids, Praunus flexuosus and Neomysis integer, and their predation on the calanoid copepod Eurytemora affinis

Community structure is shaped by external factors (i.e., habitat, temperature, and food) frequently modified by interactions among its members. This study focusses on trophic interactions between two sympatric mysids Praunus flexuosus and Neomysis integer of the Elbe Estuary, northern Germany. Based on an experimental approach, intraguild predation was evaluated. Predation rate of P. flexuosus on N. integer was positively related to predator size and temperature. Predation rate was significantly correlated with prey size, juvenile N. integer released just from the mysid marsupium being most vulnerable. However, adult P. flexuosus were able to gain more energy in terms of body carbon by catching larger N. integer, whereas immature P. flexuosus assimilated more energy by capturing large numbers of the small-sized N. integer. In contrast to N. integer, P. flexuosus showed an efficient escape behaviour that prevented all stages of N. integer from preying on any size class of P. flexuosus. When Eurytemora affinis was offered as prey, both N. integer and P. flexuosus increased predation rates with predator size and temperature. In mixed prey (N. integer and E. affinis) experiments at 10 °C, predation rates of adult P. flexuosus on N. integer released just from the marsupium declined from 17±8 to 6±4 N. integer mysid⁻¹ day⁻¹. We conclude that intraguild predation exists between the two species but is one sided with small N. integer being strongly suppressed. This heavy predation pressure is modified by the addition of alternative food resources, in this case, E. affinis.     

Seasonal variation in the mode of reproduction of Ulva intestinalis in a brackish water environment

We explored the reproductive modes of Ulva intestinalis in the inner part of the Baltic Sea during three consecutive years by using five microsatellite loci to estimate the relative abundance of diploid sporophytes and haploid gametophytes. Our results suggest that both diploid sporophytes and haploid gametophytes occur regularly in the Baltic Sea. The ratio of haploid to diploid individuals changes with seasons. Sporophytes are more abundant than gametophytes throughout the year, but the proportion of haploids increases from 10% in early summer to 35% in September. The over-wintering takes primarily place as diploid spores released by sporophytes. The sporophytes appear to reproduce both sexually and asexually in the Baltic Sea, since clones were found for this life phase. The fraction of individuals which belonged to an apparent diploid clone was higher in spring (62%) than in autumn (33%). We also found evidence for asexual clones in haploid gametophytes. The presence of both diploid and haploid individuals and the pattern of genetic and genotypic diversity provide evidence of sexual reproduction in the Baltic Sea. Thus the sporophytes and gametophytes do not function as two reproductively separate units. Compared with many other algal species with a reduced reproductive cycle in low salinity, U. intestinalis differs by having a multitude of reproductive modes also in the brackish water Baltic Sea, which can in part explain the dynamic propagation and high adaptability of the species.     

The mitochondrial genomes of Amphiascoides atopus and Schizopera knabeni (Harpacticoida: Miraciidae) reveal similarities between the copepod orders Harpacticoida and Poecilostomatoida

Members of subclass Copepoda are abundant, diverse, and—as a result of their variety of ecological roles in marine and freshwater environments—important, but their phylogenetic interrelationships are unclear. Recent studies of arthropods have used gene arrangements in the mitochondrial (mt) genome to infer phylogenies, but for copepods, only seven complete mt genomes have been published. These data revealed several within-order and few among-order similarities. To increase the data available for comparisons, we sequenced the complete mt genome (13,831 base pairs) of Amphiascoides atopus and 10,649 base pairs of the mt genome of Schizopera knabeni (both in the family Miraciidae of the order Harpacticoida). Comparison of our data to those for Tigriopus japonicus (family Harpacticidae, order Harpacticoida) revealed similarities in gene arrangement among these three species that were consistent with those found within and among families of other copepod orders. Comparison of the mt genomes of our species with those known from other copepod orders revealed the arrangement of mt genes of our Harpacticoida species to be more similar to that of Sinergasilus polycolpus (order Poecilostomatoida) than to that of T. japonicus. The similarities between S. polycolpus and our species are the first to be noted across the boundaries of copepod orders and support the possibility that mt-gene arrangement might be used to infer copepod phylogenies. We also found that our two species had extremely truncated transfer RNAs and that gene overlaps occurred much more frequently than has been reported for other copepod mt genomes.     

hunchback and Ikaros-like zinc finger genes control reproductive system development in Caenorhabditis elegans

Here we provide evidence for a C2H2 zinc finger gene family with similarity to Ikaros and hunchback. The founding member of this family is Caenorhabditis elegans ehn-3, which has important and poorly understood functions in somatic gonad development. We examined the expression and function of four additional hunchback/Ikaros-like (HIL) genes in C. elegans reproductive system development. Two genes, ehn-3 and R08E3.4, are expressed in somatic gonadal precursors (SGPs) and have overlapping functions in their development. In ehn-3; R08E3.4 double mutants, we find defects in the generation of distal tip cells, anchor cells, and spermatheca; three of the five tissues derived from the SGPs. We provide in vivo evidence that C. elegans HIL proteins have functionally distinct zinc finger domains, with specificity residing in the N-terminal set of four zinc fingers and a likely protein-protein interaction domain provided by the C-terminal pair of zinc fingers. In addition, we find that a chimeric human Ikaros protein containing the N-terminal zinc fingers of EHN-3 functions in C. elegans. Together, these results lend support to the idea that the C. elegans HIL genes and Ikaros have similar functional domains. We propose that hunchback, Ikaros, and the HIL genes arose from a common ancestor that was present prior to the divergence of protostomes and deuterostomes.