Eggs Of Daphnia Research Articles

Refining the evolutionary time machine: An assessment of whole genome amplification using single historical Daphnia eggs.

Whole genome sequencing is instrumental for the study of genome variation in natural populations, delivering important knowledge on genomic modifications and potential targets of natural selection at the population level. Large dormant eggbanks of aquatic invertebrates such as the keystone herbivore Daphnia, a microcrustacean widespread in freshwater ecosystems, provide detailed sedimentary archives to study genomic processes over centuries. To overcome the problem of limited DNA amounts in single Daphnia dormant eggs, we developed an optimized workflow for whole genome amplification (WGA), yielding sufficient amounts of DNA for downstream whole genome sequencing of individual historical eggs, including polyploid lineages. We compare two WGA kits, applied to recently produced Daphnia magna dormant eggs from laboratory cultures, and to historical dormant eggs of Daphnia pulicaria collected from Arctic lake sediment between 10 and 300years old. Resulting genome coverage breadth in most samples was ~70%, including those from >100-year-old isolates. Sequence read distribution was highly correlated among samples amplified with the same kit, but less correlated between kits. Despite this, a high percentage of genomic positions with single nucleotide polymorphisms in one or more samples (maximum of 74% between kits, and 97% within kits) were recovered at a depth required for genotyping. As a by-product of sequencing we obtained 100% coverage of the mitochondrial genomes even from the oldest isolates (~300years). The mitochondrial DNA provides an additional source for evolutionary studies of these populations. We provide an optimized workflow for WGA followed by whole genome sequencing including steps to minimize exogenous DNA.

Live observation of the oviposition process in Daphnia magna.

In favorable conditions, Daphnia magna undergoes parthenogenesis to increase progeny production in a short time. However, in unfavorable conditions, Daphnia undergoes sexual reproduction instead and produces resting eggs. Here, we report live observations of the oviposition process in Daphnia magna. We observed that the cellular contents flowed irregularly through the narrow egg canal during oviposition. Amorphous ovarian eggs developed an oval shape immediately after oviposition and, eventually, a round shape. Oviposition of resting eggs occurred in a similar way. Based on the observations, we propose that, unlike Drosophila eggs, Daphnia eggs cannot maintain cytoplasmic integrity during oviposition. We also determined that the parthenogenetic eggs were activated within 20 min, as demonstrated by vitelline envelope formation. Therefore, it is plausible that the eggs of Daphnia magna may be activated by squeezing pressure during oviposition.

Good Daphnia parents do not control the offspring microbiome.

In Focus: Mushegian, A. A., Walser, J. -C., Sullam, K. E., & Ebert, D. (2018). The microbiota of diapause: How host-microbe associations are formed after dormancy in an aquatic crustacean. Journal of Animal Ecology, 87, 400-413. https://doi.org/10.1111/1365-2656.12709. All animals are colonized by micro-organisms, most of which are benign or beneficial. Where do these micro-organisms come from? Theory predicts that micro-organisms which are transmitted vertically from parent to offspring are especially likely to be beneficial to the host, while horizontally acquired micro-organisms are opportunistic and more variable in their impact on host performance. In this issue, Mushegian etal. () investigate the source of bacteria that are required for the growth and development of Daphnia water fleas to reproductive adults. They find that, although vertically transmitted bacteria can occur in the capsule enclosing the Daphnia eggs, the micro-organisms that promote Daphnia performance are associated with the external surface of the capsule and are of likely environmental origin. This mode of transmission may be adaptive for Daphnia because, linked to the longevity and capacity for long-distance dispersal of these eggs, the environmental circumstances encountered by parent and offspring may be different; with the implication, the parental micro-organisms may not be optimal for the offspring. This study demonstrates that, although some animals require symbioses with specific coevolved, vertically transmitted microbial symbionts, other animals have evolved dependence on taxonomically variable micro-organisms of environmental origin.

Different thermal stimuli initiate hatching of Daphnia diapausing eggs originating from lakes and temporary waters

The simultaneous hatching of Daphnia resting eggs from a number of different lakes and ponds using one set of hatching cues can be difficult to achieve as environmental conditions differ considerably between water bodies. Therefore, optimal hatching conditions for ephippial eggs originating from shallow temporary waters may differ from those found in large lakes. The aim of our study was to compare the optimal thermal conditions for hatching ephippial eggs of Daphnia found in permanent lakes and those from shallow temporary ponds. We used ephippial eggs of Daphnia from the longispina species complex originating from two temporary city ponds and two deep lakes in Poland. The ephippia were protected against overheating at all stages of the field and laboratory work to prevent activation or killing of the eggs. After a refractory period (imposed storage in cool and dark conditions), ephippia were incubated at six different temperatures (6, 9, 12, 15, 18 and 21 °C) under a 16:8 L:D light regime. Our results indicate that hatching of resting eggs of Daphnia that inhabit lakes or ponds may require different thermal conditions. The hatching success of ephippial eggs originating from temporary waters was relatively high (30–56%) at all tested incubation temperatures, while for the ephippial eggs from the deep lakes it was lower (7–37%) and inversely related to water temperature. The divergent hatching responses of the ephippial eggs originating from temporary pools and lakes may reflect the typical thermal conditions during hatching in their native habitats. While in the deep lakes of the temperate zone Daphnia hatching typically occurs during the low water temperatures of early spring, in shallow ponds Daphnia hatching may occur throughout the year at varying water temperatures, from a few to over a dozen degrees Celsius.

High temperatures and absence of light affect the hatching of resting eggs of Daphnia in the tropics.

Temperature and light are acknowledged as important factors for hatching of resting eggs. The knowledge of how they affect hatching rates of this type of egg is important for the comprehension of the consequences of warming waters in recolonization of aquatic ecosystems dependent on dormant populations. This study aimed at comparing the influence of different temperature and light conditions on hatching rates of Daphnia ambigua andDaphnia laevis resting eggs from tropical environments. The ephippia were collected in the sediment of three aquatic ecosystems, in southeastern Brazil. For each lake, the resting eggs were exposed to temperatures of 20, 24, 28 and 32 °C, under light (12 h photoperiod) and dark conditions. The results showed that the absence of light and high temperatures have a negative influence on the hatching rates. Statistical differences for hatching rates were also found when comparing the studied ecosystems (ranging from 0.6 to 31%), indicating the importance of local environmental factors for diapause and maintenance of active populations.

Comparison of morphological defences in asexually and sexually reproduced eggs of Daphnia (D. galeata and D. similis) against fish kairomones

In the present study, we investigated morphological changes of hatchlings from asexually and sexually produced eggs of Daphnia galeata and D. similis against fish kairomones. In both species, hatchlings from asexual eggs showed induced traits, such as short body length, long tail spine and high relative tail spine length when influenced by fish kairomones. However, hatchlings from sexual eggs expressed high relative tail spine length regardless of kairomones presence. The results showed that the inducibility of morphological defences depends on the egg type, with inducible defences predominant in asexual eggs while fixed defences are employed by the offspring of sexual eggs in Daphnia.

Resting eggs as new biosorbent for preconcentration of trace elements in various samples prior to their determination by FAAS.

In this study, the resting eggs of aquatic creatures living in freshwater (Daphnia, Cladocera, Crustacean) ecosystems were used as a novel biosorbent extractant for synchronous preconcentration of trace Cd(II), Co(II), Cu(II), Mn(II), and Ni(II) previous to measurement by flame atomic absorpiton spectrometry (FAAS). Using column procedures, optimization studies were conducted to realize the effective adsorption of the analyte ions such as the solution pH, amount of the biosorbent, volume of the sample, interfering ions, etc. A high preconcentration factor of 67 and low relative standard deflection of ≤4.1% (n=8) were obtained. The invention constrains based on the 3 s/b criterion were 2.4 for Cd(II), 41.4 for Co(II), 4.2 for Cu(II), 3.0 for Mn(II), and 9.6 μg L(-1) for Ni(II). The accuracy of the method was verified by analysis of a certified standard reference material. The used procedure was applied to the definition of the analytes in diverse environmental samples with convincing results. Consequently, the resting eggs of Daphnia can be used as a biosorbent for preconcentration and biosorption studies.

Visualization of ecdysteroid activity using a reporter gene in the crustacean, Daphnia

Ecdysone is a hormone known to play a pivotal role in crustaceans and insects. In order to evaluate the ecdysone activities in the environment and within the organism, we have developed a biomonitoring Daphnia strain by introducing a reporter gene. In this study, the ecdysone response element was inserted in the upstream region of a reporter gene, and the DNA construct was injected into Daphnia eggs. The expression of the reporter gene was detected during the early embryonic development stage. In addition, when the eggs expressing the reporter gene were exposed to ecdysone, there was enhanced expression of the reporter gene at detectable levels, while the presence of an antagonist led to its downregulation. These results suggested that this system could be potentially developed for monitoring ecdysone activities in media.

Threshold dietary polyunsaturated fatty acid concentrations for Daphnia pulex growth and reproduction

Knowing when, where, and why food quality limits herbivore growth and reproduction is the core question in nutritional ecology. We investigated dietary threshold concentrations of polyunsaturated fatty acids (PUFAs) for the herbivorous zooplankter Daphnia pulex when fed the cyanobacterium Microcystis aeruginosa or the green algae Scenedesmus obliquus. Phosphatidylcholine liposomes were used to supplement diets with arachidonic acid (ARA, 20:4ω6), stearidonic acid (SDA, 18:4ω3), or eicosapentaenoic acid (EPA, 20:5ω3) at concentrations (per mg carbon) ranging from 0.04 to 10 μg (mg C)−1. ARA supplementation had no effect on Daphnia somatic growth or reproduction. SDA supplementation did not affect Daphnia growth, however egg production dropped below a 50% saturation threshold when dietary SDA concentrations were <0.34–0.39 μg SDA (mg C)−1. Daphnia egg production dropped below a 50% saturation threshold when EPA concentrations for the Microcystis and Scenedesmus treatments were <0.25 and 0.17 μg EPA (mg C)−1, respectively. Growth declined below the 50% threshold when EPA concentrations for the 2 diets were <0.14 and 0.04 μg EPA (mg C)−1, respectively. These results show the demands for EPA are more intense for reproduction than for somatic growth. We used the EPA results to develop an overall model between Daphnia production and dietary EPA: normalized production = (EPA/C)/(0.15 + EPA/C), R2 = 0.72, n = 40. The bootstrapped 50% saturation threshold for this model was 0.15 ± 0.03 (± SE) μg EPA (mg C)−1. Our results suggest Daphnia pulex dietary EPA limitation may be common in eutrophic lakes

High predation is of key importance for dominance of small-bodied zooplankton in warm shallow lakes: evidence from lakes, fish exclosures and surface sediments

The mean body size of limnetic cladocerans decreases from cold temperate to tropical regions, in both the northern and the southern hemisphere. This size shift has been attributed to both direct (e.g. physiological) or indirect (especially increased predation) impacts. To provide further information on the role of predation, we compiled results from several studies of subtropical Uruguayan lakes using three different approaches: (i) field observations from two lakes with contrasting fish abundance, Lakes Rivera and Rodo, (ii) fish exclusion experiments conducted in in-lake mesocosms in three lakes, and (iii) analyses of the Daphnia egg bank in the surface sediment of eighteen lakes. When fish predation pressure was low due to fish kills in Lake Rivera, large-bodied Daphnia appeared. In contrast, small-sized cladocerans were abundant in Lake Rodo, which exhibited a typical high abundance of fish. Likewise, relatively large cladocerans (e.g. Daphnia and Simocephalus) appeared in fishless mesocosms after only 2 weeks, most likely hatched from resting egg banks stored in the surface sediment, but their abundance declined again after fish stocking. Moreover, field studies showed that 9 out of 18 Uruguayan shallow lakes had resting eggs of Daphnia in their surface sediment despite that this genus was only recorded in three of the lakes in summer water samples, indicating that Daphnia might be able to build up populations at low risk of predation. Our results show that medium and large-sized zooplankton can occur in subtropical lakes when fish predation is removed. The evidence provided here collectively confirms the hypothesis that predation, rather than high-temperature induced physiological constraints, is the key factor determining the dominance of small-sized zooplankton in warm lakes.

Trehalose and Vitreous States: Desiccation Tolerance of Dormant Stages of the CrustaceansTriopsandDaphnia

Several aquatic organisms are able to withstand extreme desiccation in at least one of their life stages. This is commonly known as "anhydrobiosis." It was often thought that to tolerate such a desiccated state required high amounts of compatible solutes such as the nonreducing disaccharide trehalose, which protects cellular structures by water replacement and glass formation. Trehalose levels of dormant eggs and cysts of five freshwater crustaceans (Daphnia magna, Daphnia pulex, Triops longicaudatus, Triops cancriformis, and Triops australiensis) were observed in different states of hydration and dehydration. Although trehalose was detected in all species, the concentration was under 0.5% of the dry weight (0.05 μg/μg protein), and no change between the different states was observed. Differential scanning calorimetry (DSC) measurements indicated that dried cysts of all Triops species were in a glassy state, supporting the vitrification hypothesis. No indication for a vitreous state was found in dried resting eggs of Daphnia.

Response of diapausing eggs hatching to changes in temperature and the presence of fish kairomones

It has been hypothesized that the production of diapausing eggs in Daphnia can be induced by fish kairomones. A population of Daphnia could survive severe predation using this predator avoidance strategy. However, in changing environments, diapausing eggs experience various temperature conditions, and hatchlings at emergence may be exposed to the same predation risks as their mothers. Therefore, staying in diapause or an immediate response upon hatching to available environmental information could be important for hatchling survival. For this study, we investigated the impact of water temperature (10, 15, 20, and 25°C) in the presence and absence of fish kairomones (Lepomis macrochirus) on the hatching success of resting eggs (D. galeata). Results show that no diapausing eggs hatched at the lowest temperature (10°C), and the highest hatch percentage occurred at 15°C. Although higher water temperatures reduced hatching success, diapausing eggs hatched more quickly. The number of hatchlings was significantly higher after exposure to fish kairomones, and this was more noticeable at higher temperatures (20 and 25°C). The present results suggest that the diapausing eggs were produced as a predator avoidance strategy in Daphnia; however, the presence of fish works as a positive signal to increase hatchlings when the diapausing stage is terminated.

Maternal experience can enhance production of resting eggs in Daphnia exposed to the risk of fish predation

In many Daphnia species, hatching of resting (ephippial) eggs requires fertilization. The production of ephippial eggs can occur by individual ontogenesis, but production of males in Daphnia has to be induced by maternal experience. If ephippia appear in a population with no males present, the eggs are condemned to die. This study shows that maternal experience of predator (fi sh) presence can signifi cantly increase the tendency of females to produce ephippia in a receptive D. magna clone. The involvement of a maternal effect, not only in the production of males but also in the production of resting eggs can potentially reduce the negative effect of asynchrony between these processes.

Daphnia and ciliates: who is the prey?

The fortuitous observation of a Daphnia infested by Gymnostomatida led us to hypothesize that egg predation by ciliates could be an important mechanism regulating cladoceran population dynamics. In light of this hypothesis we reanalysed data on the dynamics of Daphnia eggs and of one Gymnostomatida species from Lago Maggiore (1996). This communication is mainly a plea to examine data where both cladocerans and Gymnostomatida were collected, to verify if the occurrence of this yet undescribed eggvoricity is widespread. Further research is needed to investigate the ecological significance of this phenomenon.

Variation in ephippial buoyancy in Daphnia pulicaria

Summary1. Freshwater zooplankton often produce diapausing eggs to survive environmental stress. The diapausing eggs of Daphnia (Crustacea, Cladocera) are encased in an ephippium that either floats at the surface or sinks to the sediment. These two types of ephippia may represent different strategies between spatial (floaters) and temporal (sinkers) dispersal of offspring.2. We observed floating and sinking characteristics of ephippia obtained from eight lakes. We then conducted an experiment with 26 Daphnia pulicaria clones obtained from six of these lakes and observed the production of buoyant and non‐buoyant ephippia under high and low food conditions.3. Ephippia were more often non‐buoyant than buoyant both from females caught in nature and those reared in the laboratory. The experiment revealed that each clone was able to produce both types of ephippia, but that there was considerable among‐clone variation in the percentage of non‐buoyant ephippia produced.4. We conclude that production of non‐buoyant versus buoyant ephippia may be driven by both genetic and environmental factors.

STRESSFUL SEX

Sex and stress may be more closely linked than you might think. Gerald LeBlanc of North Carolina State University is intrigued by the relationship between environmental stress responses and reproduction. Reproduction in the tiny freshwater crustacean Daphnia is normally an asexual all-female affair. But a stressful environment can trigger a switch to sexual reproduction in these creatures, producing freeze- and desiccation-tolerant eggs with better chances of coping with a hostile environment. Exploring this relationship between sex and stress, LeBlanc and colleagues found that the same hormonal pathway is responsible for two common environmental stress responses in Daphnia (p. 15).LeBlanc already knew that a hormone called methyl farnesoate causes Daphnia eggs to develop into males. He had also noticed that Daphnia exposed to this hormone turned a copper colour. `Then I received a serendipitous phone call from Thomas Gorr of Harvard Medical School' LeBlanc recalls, `who suggested that the animals might be turning red because the hormone was triggering production of the oxygen-transporter haemoglobin.' Puzzled by this apparent link between haemoglobin accumulation and male offspring production, LeBlanc and Gorr began a joint venture to see if the methyl farnesoate hormone was regulating both of these stress responses in the little crustacean.`If the same hormonal signalling pathway regulates haemoglobin production and male offspring production' LeBlanc reasoned, `exposing Daphnia to the hormone should trigger both haemoglobin and male offspring production.'The team decided to examine the response of 11 Daphnia clones to the hormone; five of these could produce male offspring, but the other six could not produce males. Sure enough, after the hormone treatment, haemoglobin levels in the five male-producing clones shot up and all but one of the clones also began producing male offspring. Clearly, the hormone was triggering both haemoglobin and male offspring production in these clones. But would the non-male-producing clones also increase their haemoglobin and male offspring production in response to the hormone? The team found that five of the six non-male-producing clones did not produce male offspring or haemoglobin after hormone treatment. This suggests that these clones lack a common component of the signalling pathway, most likely a receptor for the hormone. The co-responsiveness of haemoglobin accumulation and male offspring production in these 11 clones indicates that the same hormonal pathway regulates these two processes. `Discovering that haemoglobin is hormonally regulated is exciting'says LeBlanc, `because we already know a lot about the haemoglobin gene that responds to methyl farnesoate. So we now have an array of genetic tools to study this new stress signalling pathway.'But why are haemoglobin and male offspring production linked in this way?`Stressful environmental signals that tell the animals to switch to sex might coincidentally be correlated with low environmental oxygen levels' LeBlanc suggests, `for example, overcrowding may result in a drop in oxygen levels. When the environment has suddenly turned hostile, increased haemoglobin production to help the animals transport oxygen and male offspring production resulting in eggs with better survival chances might both come in handy.'

Factors influencing Gonyostomum semen blooms in a small boreal reservoir lake

Blooms of the nuisance alga Gonyostomum semen occurred in Lake 979 (Experimental Lakes Area), a small brown-water lake, that was subjected to several years of an experimental flooding regime. During periods of flooding, blooms of G. semen developed when light decreased below 100μmolm−2s−1 and total phosphorous concentrations increased to >30μgl−1. Gonyostomum semen biomass was significantly correlated with total P and DOC concentrations. In addition, G. semen abundance increased at times when Daphnia rosea had rapidly declined to<10 animals l−1. Daphnia egg ratios suggest that declines in Daphnia abundance were the result of shifts in mortality and not causally linked to changes in G. semen densities. The results observed in Lake 979 were supported by a laboratory experiment where the appearance of G. semen from Lake 979 sediment was stimulated by altering chemical and biological variables. The stimulation of G. semen blooms appears to be dependant on multiple variables rather than a single variable.

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.

Genetic versus environmental influence on pigment variation in the ephippia ofDaphnia pulicaria

Summary1. The dormant eggs ofDaphnia(Crustacea: cladocera) are encased in a protective, chitonous casing known as an ephippium. Ephippia are pigmented with melanin, and the degree of pigmentation ranges from transparent to opaque. Variation in pigmentation exists within and across populations and species, raising questions about the factors that influence the natural distribution of pigmentation.2. We used image analysis to quantify pigmentation in ephippial casings ofDaphnia pulicariathat were produced both in the field and in the laboratory. The percentage of the surface area of ephippia that is darkly pigmented ranged from 0.5 to 99.5%. The range, mean and variance of ephippial pigmentation varied across our 11 study lakes.3. Laboratory experiments compared clonal variation (five clones/lake) and population variation (five lakes) at three temperatures (15 °C, 20 °C and 25 °C). The degree of pigment variation between clones was much greater than pigment variation between temperatures, indicating that there is a genetic component to pigmentation in the ephippia which is stronger than the effect of temperature.4. Comparisons of pigment levels and lake characteristics were used to identify physical and biological factors associated with ephippial pigmentation. Investment in ephippial production was the strongest predictor with darker ephippia occurring more often in lakes with the highest ephippial production.

'Ancient' DNA in the resting egg bank of a microcrustacean can serve as a palaeolimnological database.

Recent work on the diapausing egg banks of zooplankton, such as Daphnia (Crustacea: Anomopoda), indicates that these eggs can remain viable for decades while, theoretically, DNA can remain intact for even longer periods (i.e. centuries or millennia). We isolated diapausing eggs of Daphnia from a 30 m long sediment core taken from a hypereutrophic, northern German lake (Belauer See), with some eggs found in dated core material as old as 4500 years. Using microsatellite markers, we analysed the genetic structure of the resting eggs dated as old as ca. 200 years, and found that, although levels of heterozygosity remained remarkably stable, significant genetic differentiation (Nei's D = 0.36; F(ST) = 0.15) between recent and 'ancient' resting eggs (including allele frequency shifts and private alleles) was detected. These shifts represent either species-level changes in this complex (i.e. species-specific characters of ephippia are not always robust), or intraspecific shifts in genetic variation, or a combination of both. This study demonstrates that the egg banks of aquatic zooplankton can serve as repositories of both genetic (intrapopulational) and ecological (interspecific) information. The use of molecular markers, such as microsatellites, on diapausing egg/seed banks may open new avenues of enquiry related to tracking the long-term genetic (and/or species) shifts that are associated with long-term environmental changes.