Magnetic characterization of Daphnia resting eggs

This study characterized the magnetic materials found within Daphnia resting eggs by measuring static magnetization with a superconducting quantum interference device (SQUID) magnetometer, after forming two types of conditions, each of which consists of zero-field cooling (ZFC) and field cooling (FC). Magnetic ions, such as Fe(3+), contained in Daphnia resting eggs existed as (1) paramagnetic and superparamagnetic particles, demonstrated by a magnetization and temperature dependence of the magnetic moments under an applied magnetic field after ZFC and FC, and (2) ferromagnetic particles with definite magnetic moments, the content of which was estimated to be very low, demonstrated by the Moskowitz test. Conventionally, biomagnets have been directly detected by transmission electron microscopes (TEM). As demonstrated in this study, it is possible to nondestructively detect small biomagnets by magnetization measurement, especially after two types of ZFC and FC. This nondestructive method can be applied in detecting biomagnets in complex biological organisms.     

Methods for the activation of the resting eggs of Daphnia

SUMMARY. 1. The conditions required to initiate development of resting eggs of thirty-six clones of Daphnia representing seven species were investigated.
2. The temperature of both dark incubation and subsequent light treatment are shown to affect hatch success. By varying these parameters the majority of resting eggs from each test clone were stimulated to develop. Arctic clones required a low hatching temperature (7°C), whereas clones from warmer climates hatched best at 14–21°C.
3. Variation in hatching cues existed between conspecific individuals from different collection sites. These differences suggest that research determining macro- and microgeographic patterns in hatching phenology would be fruitful.     

Elemental characterization of Daphnia resting eggs by X-ray analytical microscopy

Resting eggs of Daphnia, a key crustacean zooplankton of freshwater food chains, can remain viable for more than a century. These eggs are able to withstand freezing and drying, and can survive the harsh environment of a predator's digestive system. Until recently little was known about the chemical composition, microanatomy, and physical properties of the resting eggs. The current study utilized a physical technique, the X-ray analytical microscope, to identify and localize component elements of the Daphnia resting egg. The analysis demonstrated that phosphorus, sulfur, potassium, and calcium were detected as elemental components of the resting egg, and detection intensities of the four elements differed according to the position of the eggs. Phosphorus and calcium were mostly detected in regions of the eggshell that surrounded the two embryos. In addition, sulfur was distributed throughout the eggshell whereas potassium was localized to the areas that corresponded to where the embryos were encased. Through the use of X-ray analytical microscopy, the current study identifies elemental characteristics in relationship to the structure of the Daphnia resting eggs.     

Size-selective dispersal of Daphnia resting eggs by backswimmers (Notonecta maculata)

Freshwater zooplankton is increasingly used to study effects of dispersal on community and metacommunity structure. Yet, it remains unclear how zooplankton disperses. Clearly, birds and wind play a significant role as zooplankton dispersal agents, but they may not always be the main vectors. This experimental study shows that a cosmopolitan aquatic insect, Notonecta, can be an important vector of cladoceran resting eggs (ephippia). Dispersing Notonecta frequently transported ephippia during flight, with a bias towards smaller ephippia in two species. A similar trend was present at the species level: Daphnia species with smaller ephippia were more often dispersed, suggesting that Notonecta could generate specific colonist communities. In addition, buoyancy appeared a critical trait, as non-floating ephippia of Daphnia magna were never dispersed. Our data suggest that Notonecta could be important dispersers of Daphnia, and that knowledge of dispersal dynamics of Notonecta may be used to predict Daphnia dispersal, colonization and resilience to disturbance.     

Resilience to changes in lake trophic state: Nutrient allocation into Daphnia resting eggs

During past decades, many lakes underwent drastic human‐caused changes in trophic state with strong implications for population dynamics and food web processes. We investigated the influence of trophic state on nutrient allocation into Daphnia resting eggs. The production of resting eggs is an important survival strategy, allowing Daphnia to cope with unfavorable environmental conditions. Allocation of essential nutrients into resting eggs may crucially influence embryonic development and offspring survival and thus is of great ecological and evolutionary interest. The capacity of Daphnia to adjust the allocation of nutrients into resting eggs may depend on the dietary nutrient supply, which may vary with trophic state‐related changes in the phytoplankton community composition. Resting eggs were isolated from sediment cores taken from Lake Constance, a large prealpine lake with a distinct eutrophication and reoligotrophication history, and analyzed for elemental (carbon, nitrogen, and phosphorus) and biochemical (sterols and fatty acids) nutrients. Carbon allocation into Daphnia resting eggs continuously decreased over time, irrespective of changes in trophic state. The allocation of nitrogen into Daphnia resting eggs followed the changes in trophic state, that is, nitrogen concentrations in resting eggs increased with eutrophication and decreased again with reoligotrophication. The allocation of phosphorus, sterols and long‐chain polyunsaturated fatty acids, such as eicosapentaenoic acid, into Daphnia resting eggs did not change significantly over time. Changes in trophic state strikingly influenced all trophic levels in Lake Constance. However, nutrient allocation into Daphnia resting eggs was mostly resilient to changes in lake trophic state.     

The importance of hatching from resting eggs for population dynamics and genetic composition of Daphnia in a deep reservoir

1. Cyclical parthenogens, organisms switching between sexual and asexual mode of reproduction, are common in aquatic habitats. In permanent waterbodies with year‐round persistence of populations, the relevance of sexually produced resting eggs for population dynamics and structure is still largely unexplored. 2. The aim of this study was to investigate the importance of recruitment from resting eggs to Daphnia spp. in a deep dimictic reservoir with overwintering populations (Saidenbach Reservoir, Germany). We linked field studies on Daphnia population dynamics, hatching rates and diapausing egg abundances in the sediment with allozyme analysis of the genetic variability of both the pelagic population and hatchlings from resting eggs. 3. We found higher hatching rates at intermediate depths (10–18 m), although highest diapausing egg abundances were found in the deeper parts of the reservoir. From the extensive field data, we calculated a population budget for the main basin of the reservoir, which showed that hatched daphnids accounted for <1% of the total abundance and explained only a tiny fraction of population growth. 4. The genetic composition (MultiLocus Genotypes, MLG) of the pelagic population was relatively stable throughout the stratified period with up to five among 39 detected MLGs attaining frequencies >5% per date. From 11 MLGs differentiated among hatched daphnids, three were recovered in the pelagic population in perceptible, albeit low frequencies (1–4%). 5. We conclude that the contribution of hatched animals to the spring population dynamics is negligible in situations with an overwintering population present and will only become relevant if densities are extremely low. In this type of system, the significance of the resting egg bank is presumably mainly related to the maintenance of genetic diversity.     

1,25-Dihydroxyvitamin D3 injections into rat fetuses : effects on fetal plasma calcium, plasma phosphate and mineral content

The in vivo effects of 1,25-(OH)2D3 were assessed using fetuses from normal and thyroparathyroidectomized (TPTX) pregnant rats. 21.5-day old decapitated fetuses from TPTX mothers exhibited lowered basal plasma calcium, elevated basal plasma phosphate and an increased percentage of total ash compared to intact littermates. In decapitated fetuses from normal mothers, neither plasma calcium nor plasma phosphate was changed. Subcutaneous injection of 1 micrograms of 1,25-(OH)2D3/kg of body weight to 19.5-day old fetuses (intact or deprived of their parathyroid glands by decapitation) from TPTX mothers induced a marked rise in plasma calcium levels (2.01 and 3.66 mg/dl, respectively) 48 h later. Little change occurred in fetuses from normal mothers (1.06 mg/dl in decapitated and no change in intacts). A decrease in plasma phosphate levels was observed with the same dose in both decapitated and intact fetuses from TPTX mothers (- 1.39 and - 0.65 mg/dl, respectively), while no modification was found in fetuses from normal females. Therefore, the hypersensitivity of fetuses from TPTX mothers to 1,25-(OH)2D3 was unrelated to the development of the fetal hyperparathyroidism secondary to maternal TPTX. The percentage of ash was unchanged in decapitated fetuses from TPTX mothers and was increased in intact littermates after 1,25-(OH)2D3 treatment. However, these values for total ash may represent alterations in bones and/or soft tissues.     

萼花臂尾轮虫休眠卵萌发的研究

报道了萼花臂尾轮虫休眠卵的低温贮存时间、萌发温度、光照、孵化液等因子对其萌发的影响．休眠卵萌发率随低温贮存时间的延长而提高,以贮存30～50d作用最显著．在10～30℃温度范围内．萌发率随温度的升高而增加,超过30℃时,萌发率则有所下降．光对休眠卵的萌发是一非必需条件。把休眠卵从不新鲜的孵化液转移到新鲜的孵化液中能刺激其萌发．     

Solid-state phosphorus-31 nuclear magnetic resonance studies of synthetic solid phases of calcium phosphate: potential models of bone mineral

Phosphorus-31 NMR spectra have been obtained from a variety of synthetic, solid calcium phosphate mineral phases by magic angle sample spinning. The samples include crystalline hydroxyapatite, two type B carbonatoapatites containing 3.2 and 14.5% CO3(2-), respectively, a hydroxyapatite in which approximately 12% of the phosphate groups are present as HPO4(2-), an amorphous calcium phosphate, monetite, brushite, and octacalcium phosphate. Spectra were observed by the standard Bloch decay and cross-polarization techniques, as well as by a dipolar suppression sequence, in order to distinguish between protonated and unprotonated phosphate moieties. The spectra of the synthetic calcium phosphates provide basic information that is essential for interpreting similar spectra obtained from bone and other calcified tissues.     

Whole genome amplification and sequencing of a Daphnia resting egg

Resting eggs banks are unique windows that allow us to directly observe shifts in population genetics, and phenotypes over time as natural populations evolve. Though a variety of planktonic organisms also produce resting stages, the keystone freshwater consumer, Daphnia, is a well‐known model for paleogenetics and resurrection ecology. Nevertheless, paleogenomic investigations are limited largely because resting eggs do not contain enough DNA for genomic sequencing. In fact, genomic studies even on extant populations include a laborious preparatory phase of batch culturing dozens of individuals to generate sufficient genomic DNA. Here, we furnish a protocol to generate whole genomes of single ephippial (resting) eggs and single daphniids. Whole genomes of single ephippial eggs and single adults were amplified using Qiagen REPLI‐g Single Cell kit reaction, followed by NEBNext Ultra DNA Library Prep Kit for library construction and Illumina sequencing. We compared the quality of the single‐egg and single‐individual amplified genomes to the standard batch genomic DNA extraction in the absence of genome amplification. At mean 20× depth, coverage was essentially identical for the amplified single individual relative to the unamplified batch extracted genome (>90% of the genome was covered and callable). Finally, while amplification resulted in the slight loss of heterozygosity for the amplified genomes, estimates were largely comparable and illustrate the utility and limitations of this approach in estimating population genetic parameters over long periods of time in natural populations of Daphnia and also other small species known to produce resting stages.     

Hatching of cladoceran resting eggs: temperature and photoperiod

Summary 1. We identified temperature and photoperiod conditions under which the hatching of 45 cladoceran species could be elicited. Identification of appropriate hatching cues is of primary importance for the exploration of the ties between active and diapausing stages.
2. Incubation temperature affected the hatching success of resting eggs isolated from Danish, Belgian/Dutch and Spanish sediments. In general, most hatchlings and species were retrieved at 15 °C. Danish and Belgian/Dutch resting eggs hatched more successfully under a long day photoperiod than in continuous illumination.
3. Most species could be retrieved after incubation of resting eggs isolated from a limited amount of sediment (0.4 kg) under a single, well chosen combination of temperature and photoperiod. Processing additional sediment samples under seven more incubation regimes only allowed detection of 21% (Spain) to 34% (Denmark) more species.
4. The incubation period for resting eggs to hatch was strongly influenced by incubation temperature. Our results show that hatching experiments aimed at assessing cladoceran species richness and conducted at 15 °C should be continued for a period of at least 2 weeks, after which a random subset of hatchlings (e.g. n  = 100) can be selected from the total hatchling assemblage.     

Energy content of spider eggs

Summary The energy content of spider eggs was determined on samples from 12 species representing 6 families. These values ranged from 26.32 to 29.00 joules per mg ash-free dry mass with a mean of 27.30±SE of 0.27. The higher values were found in those species that overwinter as developmental stages within the egg sac. Rates of energy expenditure of developing eggs and spiderlings within the egg sac were only 7 to 19% of those of emerged spiderlings. The energy conserved by this reduction in rate of metabolism may facilitate survival without feeding during the potentially long periods of aerial dispersal by ballooning, a characteristic activity of most newly emerged spiderlings. The variation in mass-specific energy content is less than variation in clutch size and individual egg size on an intra-and interspecific basis. There was no correlation between energy content per unit egg mass and size of the female parent, egg size, or clutch size. Further analysis indicated that no single measure such as clutch size accurately represents the proportional amount of energy invested in reproduction in these animals.     

New chitin,chitosan, and O-carboxymethyl chitosan sources from resting eggs of Daphnia longispina (Crustacea); with physicochemical characterization,and antimicrobial and antioxidant activities

The paper describes the isolation and characterization of chitin and chitosan from Daphnia longispina resting eggs harvested from a reservoir. Resting eggs are fertilized eggs that are encased in chitinous shells called ‘ephippia’ and which ensure the survival of the Daphnia population in adverse conditions. The chitin-content of D. longispina resting eggs was found to be 23 ～ 25% and the chitosan (having a 70 ～ 75% deacetylation degree) yield of the chitin was 76 ～ 77%. This high chitin-content indicates that D. longispina resting eggs can be exploited as a chitin source. The structure and thermal properties of chitin, extracted from D. longispina resting eggs, were characterized by employing Fourier transform infrared spectroscopy, thermogravimetric analysis, X-ray diffraction and scanning electron microscopy. The crystallinity of the chitin was found to be very low (48%). Physicochemicallycharacterized chitosan and the produced O-carboxymethyl chitosan were tested for their antimicrobial and antioxidant activity. It has been observed that chitosan displays antimicrobial activity against all pathogenic bacteria, whereas O-carboxymethyl chitosan only exhibits inhibition activity against L. garvieae, L. Monocytogenes ATCC 7644, Y. enterocolitica NCTC 11175 and S. aureus ATCC 25923. In a free radical scavenging activity assay, the IC₅₀ values of chitosan, O-carboxymethyl chitosan and butylated hydroxytoluene were found to be 23.01, 56.43 and 0.05, respectively. The ferric-reducing power of O-carboxymethyl chitosan (EC₅₀ = 8.30) indicated higher activity than chitosan (EC₅₀ = 10.12).     

The effect of embryonic development on metal and calcium content in eggs and eggshells in a small passerine

Published information relating to changes in the chemical element content of avian eggs caused by embryonic development is extremely scarce, although it may be crucial for understanding both the presence of anthropogenic pollutants as well as physiological levels of micronutrients. We assessed the variation in concentrations of calcium (Ca) and magnesium (Mg) and nine trace elements: seven essential (chromium (Cr), copper (Cu), nickel (Ni), manganese (Mn), iron (Fe), cobalt (Co) and zinc (Zn)) and two non‐essential (lead (Pb) and cadmium (Cd)) in shells and contents (both egg yolk and egg white) of embryonated and non‐embryonated eggs. We investigated the eggs of the Eurasian Reed Warbler Acrocephalus scirpaceus, a large proportion of whose eggs are infertile in our study population (almost 43% of clutches contain unhatched eggs) as well as significant embryo‐induced eggshell thinning at the equator of embryonated eggs. We found significantly higher concentrations (≥ 22.7%) of all the focal elements in the contents of embryonated eggs in comparison with non‐embryonated eggs, and a very pronounced one for Ca (nearly twice as high). The shells of embryonated eggs contained significantly higher concentrations of Zn (104.1%), Fe (56.5%), Pb (32.8%) and Cu (28.0%) but significantly lower ones of Co (8.9%) and Ca (9.3%) than the shells of non‐embryonated eggs. The simultaneous higher concentrations of all elements in the content of thinner‐shelled embryonated eggs suggest the parallel transfer of these elements along with Ca resorption from the shell into the egg interior during embryo formation. The higher concentration of most elements in the thinner shells of embryonated eggs may be indicative of the maternal deposition of some of these elements in a shell layer not subject to embryonic depletion, or in the eggshell membrane. Our results highlight the need for the careful selection of egg samples, which should differentiate between embryonated and non‐embryonated eggs in the analytical treatment of eggs and eggshells.