Maternal Effects of Photoperiod and Food Level on Life History Characteristics of the Cladoceran Daphnia Pulicaria Forbes

The response of various life-history characteristics of Daphnia pulicaria to photoperiod and food concentration was measured in 16 combinations of maternal and offspring environments (long vs. short day, high vs. low food) in flow-through experiments. Response variables in offspring were time and survival to release of first offspring, clutch size and neonate mass in the first brood, mass of adult females after 30days and somatic growth rate during the course of the experiment. Most of these parameters were directly controlled by food concentration in the offspring environment, but maternal effects frequently modified the response. A long day length in the maternal environment resulted in a prolongation of the time to first clutch release in offspring similar to the direct effect of low food. Likewise, survival to maturation and female mass were affected by maternal photoperiod. Somatic growth rate and clutch size responded to combined effects of maternal food conditions and photoperiod. The laboratory results were used to predict the seasonal change of fecundity of Daphnia in the field. When data on clutch size are ordered in a sequence as the different combinations of maternal and offspring environment occur during the seasonal succession in a temperate lake, they show a bimodal distribution with a high peak in spring and a lower peak in fall. This pattern is consistent with field observations. We conclude that photoperiod and maternal effects are important factors influencing life history and population dynamics of Daphnia.     

Life history strategies of cladocerans: comparisons of tropical and temperate taxa

We review recent works on different life history variables of cladoceran taxa in tropical and temperate freshwater bodies, comparing the strategies that cladocerans have evolved to adapt to contrasting environmental conditions in the two geographical regions. These life-history parameters relate to age and size at maturity, survival, fecundity, life-expectancy at birth, lifespan, gross, and net reproductive rates, generation time, the rate of population increase, peak population density and day of peak abundance. We also discuss the role of photoperiod and temperature on some of these life history parameters. We found a general paucity of experimental work and field data in tropics on cladocerans. There is very limited information on the few Daphnia species found in the tropics. The misconception of low species diversity of cladocerans in the tropics arose due to several reasons including lack of extensive and intensive field collections. Higher water temperatures apparently promote permanent infestation of tropical waters with toxic cyanobacteria, which reduce the zooplankton diversity. In addition to higher temperatures in the tropics, the year-round high predation pressure of planktivorous fish probably causes the tropical species, particularly in pelagic habitats, to reach maturity earlier (< 3 days) than in temperate regions. Species of Daphnia in temperate regions are particularly adapted to living at food concentrations that are much lower and seasonably more variable than those for tropical genera such as Diaphanosoma. This is further corroborated by the more than an order of magnitude higher threshold food concentration (TFC) for tropical Cladocera than for their temperate counterparts. Fecundity patterns differ between tropical and temperate cladoceran taxa: cultured under optimal temperature regimes, tropical taxa have fewer eggs than temperate species of a comparable body size. Predation pressure may act differently depending on the size of the cladoceran neonates and thus on their population size structure. Global warming and climate changes seem to affect the behaviour (migration), distribution, and abundance of cladocerans. Apparently, in direct response to these changes, the possibility of encountering the tropical cladocerans in the northern, temperate hemisphere (bioinvasions) is on the rise.     

Offspring size–number tradeoffs and food quality feedbacks impact population dynamics in a Daphnia–algae system

Population fluctuations can be affected by both extrinsic (e.g. weather patterns, food availability) and intrinsic (e.g. life‐history) factors. A key life‐history tradeoff is the production of offspring size versus number, ranging from many small offspring to few large offspring. Models show that this life‐history tradeoff in offspring size and number, through maturation time, can have significant impacts on population dynamics. However, few manipulative experiments have been conducted that can isolate life‐history effects from impacts of extrinsic factors in consumer–resource systems. We experimentally tested the effect of an offspring size–number tradeoff on population stability and food availability in a consumer–resource system. Using Daphnia pulex, we created a shift from many, small offspring being produced to fewer, larger offspring. Two sets of experiments were performed to examine the interaction of an extrinsic factor (light levels) and intrinsic population structure on dynamics, and we controlled for the ingestion pressure on algal prey at the time of the manipulation. We predicted that the tradeoff would impact internal consumer population characteristics, including biasing the stage structure towards adults, increasing adult size, and increasing average population‐level reproduction. This adult‐dominated stage structure was predicted to then lead to instability and a low quantity–high quality food state. Under all light levels, the manipulated populations became dominated by large adults. Contrary to predictions, the amplitudes of fluctuations in Daphnia biomass were lower in populations shifted to few–large offspring, representing higher stability in these populations. Furthermore, in high light conditions, a stable low Daphnia – high algae biomass (low food quality) state was observed in few–large offspring treatments but not in control (many–small offspring) treatments. Our results show a strong link between light levels as an extrinsic factor and the life‐history tradeoff of consumer offspring size versus number that impacts consumer–resource population dynamics through feedbacks with resource quality.     

Embryogenesis plasticity and the transmission of maternal effects in Daphnia pulex

Understanding how genetic, nongenetic, and environmental cues are integrated during development may be critical in understanding if, and how, organisms will respond to rapid environmental change. Normally, only post‐embryonic studies are possible. But in this study, we developed a real‐time, high‐throughput confocal microscope assay that allowed us to link Daphnia embryogenesis to offspring life history variation at the individual level. Our assay identified eight clear developmental phenotypes linked by seven developmental stages, the duration of which were correlated with the expression of specific offspring life history traits. Daphnia embryogenesis varied not only between clones reared in the same environment, but also within a single clone when mothers were of different ages or reared in different food environments. Our results support the hypothesis that Daphnia embryogenesis is plastic and can be altered by changes in maternal state or maternal environment. As well as furthering our understanding of the mechanisms underpinning parental effects, our assay may also have an industrial application if it can be used as a rapid ecotoxicological prescreen for testing the effect that pollutant doses have on offspring life histories traditionally assayed with a 21‐day Daphnia reproduction test.     

Growth rate of <Emphasis Type="Italic">Daphnia</Emphasis> feeding on seston in a Siberian reservoir: the role of essential fatty acid

Specific growth rates (based on biomass increment in unit time) of Daphnia fed natural reservoir seston from a eutrophic Siberian reservoir were studied during four vegetation seasons in a laboratory flow-through system. Concentrations of particulate C, N, and P in reservoir seston <115 μm were comparatively high, 1.9, 0.2, and 0.05 mg l⁻¹, respectively; maximum C:P ratio was 259 mol:mol. According to conventional thresholds, concentrations of elements and their stoichiometry did not limit the growth rate of Daphnia. Eicosapentaenoic acid (EPA) in seston significantly effected the growth rates at concentrations up to ∼13 μg l⁻¹ (regression ANOVAR F-test value was 7.91), but not above this concentration. Thus, we consider this concentration of EPA in seston <115 μm to be the limiting concentration of EPA for the growth of Daphnia (longispina group), i.e., below this concentration EPA was the best single predictor of the growth of Daphnia.     

Plant regeneration via somatic embryogenesis of <Emphasis Type="Italic">Camptotheca acuminata</Emphasis> in temporary immersion system (TIS)

The present study describes a protocol for plant regeneration via somatic embryogenesis in temporary immersion system (TIS) for Camptotheca acuminata. Somatic embryos were induced by culturing hypocotyl segments from 14-day-old in vitro grown C. acuminata seedlings in TIS. Hypocotyl segments were placed in culture vessels modified with a mechanical device to support the fixation of explants. Cultures were maintained under a 16 h photoperiod with a light intensity of 60 μmol m⁻² s⁻¹ PPF at 25 ± 1°C. After 16 weeks of incubation embryogenic calli were formed above the edge of the mechanical device in the basal Murashige and Skoog (MS) medium containing 35 g l⁻¹ sucrose and without hormonal supplementation. For plantlet regeneration, somatic embryos at cotyledonary stage were cultured in three different concentrations of 6-benzylamino-purine (0.5, 1.0 and 1.5 mg l⁻¹ BAP) and in plant growth regulator (PGR) free medium. In general, 0.5 mg l⁻¹ BAP was found to be the most effective concentration for growth and development of Camptotheca embryos in TIS. Conversion of somatic embryos into plantlets was also successfully achieved on sterile substrates moistened with 0.5 mg l⁻¹ BAP. Plantlets derived from cotyledonary embryos were rooted in vitro with 0.5 mg l⁻¹ indole-3-butyric acid (IBA) before transfer to ex vitro conditions.     

Effects of crowding and different food levels on growth and reproductive investment of Daphnia

The effects of daphniid crowding on juvenile growth rate, length at first reproduction, clutch size and egg size of four species of Daphnia were compared with the effects of food level. Juvenile Daphnia were grown to primipary in a flow-through system in water conditioned by different densities of the same, or another, species. At high ambient food levels, water from Daphnia that had been crowded at densities 150 l^–1 depressed growth rate and lowered body size and clutch size of D. hyalina and D. galeata; effects on the same traits of D. magna and D. pulicaria were variable (stimulation, depression, or no effect). D. hyalina and D. galeata responded to gradients of increasing daphniid density (0–300 l^–1) by altering egg mass, somatic mass and clutch size to maintain a relatively constant reproductive investment; egg mass increased with crowding and then decreased in a pattern consistent with Glazier's (1992) hypothetical model of changes in offspring size in relation to food quantity and maternal demand. Effects of crowding by conspecifics were indistinguishable from those of other species. This study, which uncouples the effect of crowding per se from ambient resource depletion, shows that chemical substances released by high densities of Daphnia can cause changes in life-history traits comparable to those that occur in response to low food levels.     

Ontogenetic Changes in Daphnia Responsiveness to Fish Kairomone

Exposing Daphnia to the presence of a predator at different stages of ontogenesis leads to different reactions. Predator-induced changes of life history parameters related to the first reproduction schedule, as age and size at first reproduction, take place in the stage preceding that one in which vitellogenesis occurs. Determination of the amount of energy allocated to this first reproduction, including offspring number, occurs in the first 24-h of life. In case of parameters related to reproduction schedule, time since induction until the release of the first clutch of offspring is tens of hours; in case of allocation patterns – more than 100. This suggests that Daphnia do not remember the information of a potential threat but, instead, the decisions on future development are undertaken once they have estimated the reigning predation regime. Shortening of the period during which eggs are carried in the brood chamber, thus shortening the time when Daphnia are more easily visible, may be another adaptation to reduce danger from fish predation and so increasing the probability of survival until the first successful reproduction.     

Influence of diet and photoperiod on development and reproduction of European populations of Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae)

The current study examines the effect of photoperiod (16:08 or 12:12 h L:D) and diet (eggs of Ephestia kuehniella Zeller (Lepidoptera: Phycitidae) or the pea aphid Acyrthosiphon pisum (Harris) (Hemiptera: Aphididae)) on the development and reproduction of the multicoloured Asian lady beetle Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae). A long-term laboratory population of H. axyridis (since 1998) and a melanic and non-melanic population originating from field collected individuals of H. axyridis in Belgium were used in this study. Long day conditions (16 h photoperiod) shortened development of the field populations with 2–3 days when compared with short day conditions (12 h photoperiod). Oviposition in the field populations was delayed by 1–3 months when reared at a 12 h photoperiod. Dissections indicated that the females were in reproductive diapause. As compared with live pea aphids, a diet consisting of E. kuehniella eggs yielded heavier adult body weights (up to 12%) and increased the number of egg laying days (by 45–169%) for both field populations at a 16 h photoperiod and lengthened adult life span (by 45–92%) under both light regimens. The morph types differed in their response to the foods offered in terms of developmental rate, pre-oviposition period and number of oviposition days. The laboratory and field strains responded differentially to regimens of food and photoperiod. The study indicated a greater nutritional plasticity of the non-melanic morphs which may offer them a competitive advantage that may in part explain the predominance of non-melanic morphs in newly colonized areas.     

The influence of <Emphasis Type="Italic">Myriophyllum verticillatum</Emphasis> and artificial plants on some life history parameters of <Emphasis Type="Italic">Daphnia magna</Emphasis>

Submerged macrophytes are known to serve as refuge for zooplankton but also seem to suppress the zooplankton growth. Thus, there is a conflict between the positive and negative role of macrophytes for zooplankton. We tested the influence of physical structure using artificial macrophytes, chemicals released by a macrophyte (Myriophyllum verticillatum), as well as the combined effects of these two factors on the life history of Daphnia magna. Daphnids matured at a smaller size and produced fewer eggs but larger individual offspring in the presence of artificial and real plants. Thus, under conditions with plants, we observed a trade off between number of eggs produced and the individual size of the offspring. Daphnids grown in the presence of exudates without plants were larger at maturity and showed no reduction in clutch size as compared with the control. We suggest that the macrophytes (real and artificial) negatively affected the daphnids in two ways: (1) food particles were settling down faster on the plant structures and this reduced the available food for Daphnia, (2) the plant structures were obstacles for the daphnids causing them to spent more energy during swimming. Both effects resulted in a reduced somatic growth, whereas the increased individual offspring size was probably a response to the reduced food level. Daphnia was not negatively affected by Myriophyllum exudates.     

An improved micropropagation of <Emphasis Type="Italic">Spilanthes acmella</Emphasis> L. through transverse thin cell layer culture

An efficient and improved in vitro propagation system for Spilanthes acmella L. using transverse thin cell layer (tTCL) culture system was established. The frequency of shoot regeneration from tTCL nodal segments was affected by concentrations of plant growth regulators and orientation of the explant. MS (Murashige and Skoog in Physiol Plant 15:473–497, 1962) medium with 5.0 mg dm⁻³ BAP was optimal for shoot regeneration. Upon this medium, the explant inoculated in the upright orientation exhibited a high frequency of shoot regeneration (about 97%), and the highest number of shoots (31.5) per explant. The intact node (1.0–1.5 cm) cultured on the same medium had significantly lower shoot multiplication ability with only 4.5 shoots per responsive explant. As compared to BAP alone, the combination of BAP and Kin or NAA did not have positive effects on shoot multiplication from tTCL nodal segments. Rooting of shoots was achieved on growth regulator free full-strength MS medium. Plantlets were transplanted into soil with 90–100% survival rate.     

Depth selection and life history strategies as mutually exclusive responses to risk of fish predation in <Emphasis Type="Italic">Daphnia</Emphasis>

We tested if pelagic crustaceans of the genus Daphnia use different anti-predator defences in environmental conditions that do or do not offer deep refuge from planktivorous fish. We kept Daphnia catawba in two series of 9-m deep enclosures with and without caged cyprinid fish Phoxinus eos. In one series of enclosures, Daphnia could select its depth of residence and hide in deep dark water layers to avoid anticipated fish predation, while in another series of enclosures, a plankton net barrier fixed at 2-m depth forced them to stay in subsurface zone exposed to fish kairomones. We compared depth residence and migratory behaviour strategies with life history strategies (body size and size at first reproduction, diapause induction) in Daphnia exposed or not to fish kairomones with or without deep refuge. In deep enclosures with fish, Daphnia spent daytime hours in deep dark layers while at night, they resided closer to the water surface. Yet, no change in life history parameters of migrating individuals was observed compared to the fish-free conditions. In enclosures with fish, where the net barrier forced Daphnia to reside in subsurface zone, they produced smaller offspring, matured at smaller size and achieved lower maximum body length compared to the fish-free conditions. However, they did not produce diapausing eggs. Our experimental study supports the hypothesis that diel vertical migration behaviour with daytime residence in deep, dark water are the preferred antipredator strategy chosen by Daphnia facing anticipated fish predation over life history changes such as reduced size and low growth rate which are used when dark deep refuge is not present or accessible.     

Life history consequences of sterol availability in the aquatic keystone species <Emphasis Type="Italic">Daphnia</Emphasis>

The absence of essential biochemical nutrients, such as polyunsaturated fatty acids or sterols, has been considered as a mechanism determining trophic interactions between the herbivore Daphnia and its phytoplankton food source. Here, we experimentally quantify the sensitivity of two Daphnia species to decreasing amounts of dietary sterols by measuring variations in life history traits. The two species Daphnia magna and D. galeata were fed different mixtures of the sterol-containing green alga Scenedesmus obliquus and the sterol-free cyanobacterium Synechococcus elongatus; a higher proportion of Synechococcus in the food is equivalent to a decrease in dietary sterols. To address the significance of sterol limitation, the Daphnia species were also fed Synechococcus supplemented with cholesterol. In both species, somatic and population growth rates, maternal dry mass, the number of viable offspring, and the probability of survival were significantly reduced with the lower availability of sterols. A high correlation between the sterol content of the mixed diet and the somatic and population growth rates was found, and growth on cholesterol-supplemented Synechococcus fitted well into this correlation. Somatic growth of first-clutch neonates grown on 100% Synechococcus exhibited a pattern similar to that of somatic growth of their mothers grown on the different food regimes, which demonstrated the significance of maternal effects for sterol-limited population growth. Daphnia galeata had a twofold higher incipient limiting sterol level than D. magna, which indicated interspecific differences in sterol requirements between the two Daphnia species. The results suggest a strong impact of dietary sterols on life history traits and therefore, population dynamics of the keystone species Daphnia.     

Sensitivity of Daphnia species to phosphorus-deficient diets

The life history of freshwater cladocerans such as Daphnia spp. is strongly affected by their environment. Factors such as temperature, food quantity and even the presence or absence of predators influence growth, reproduction and morphology of individuals. Recently, it has also become clear that the quality of the food can affect various life history traits of Daphnia. More specifically, the effect of the elemental composition of algae, expressed as the C:P ratio, has been studied intensively. Daphnia species differ in their response to differences in the C:P ratio of their food. Until now, it has been unclear whether these species differences are driven by phylogenetic constraints or by adaptation to particular environmental conditions. Here we present laboratory experiments with 12 Daphnia species from three different subgenera originating from a broad range of habitats. We compared somatic growth rates and sensitivity to variation in the nutrient stoichiometry of the food with habitat parameters, taking into account the phylogenetic history of the species. No associations between fitness and habitat parameters were detected. However, we found a trade-off between sensitivity to P-deficient diets and the maximum growth rate on a P-sufficient diet. In several cases, this trade-off helps to explain the association between species distribution and habitat parameters. We observed no correlation of the sensitivity to P limitation with the phylogenetic history of the genus Daphnia. Thus, we conclude that the differential responses among Daphnia species to variation in P content in food were driven mainly by adaptations to their local habitats, and are not constrained by deep evolutionary patterns.     

Nutrient enrichment in the High Arctic associated with Thule Inuit whalers: a paleolimnological investigation from Ellesmere Island (Nunavut,Canada)

The combined influence of a pesticide (carbaryl) and a cyanotoxin (microcystin LR) on the life history of Daphnia pulicaria was investigated. At the beginning of the experiments animals were pulse exposed to carbaryl for 24 h and microcystins were delivered bound in Microcystis’ cells at different, sub-lethal concentrations (chronic exposure). In order to determine the actual carbaryl concentrations in the water LC–MS/MS was used. For analyses of the cyanotoxin concentration in Daphnia’s body enzyme-linked immunosorbent assay (ELISA) was used. Individual daphnids were cultured in a flow-through system under constant light (16 h of light: 8 h of dark), temperature (20°C), and food conditions (Scenedesmus obliquus, 1 mg of C l⁻¹). The results showed that in the treatments with carbaryl egg numbers per female did not differ significantly from controls, but the mortality of newborns increased significantly. Increasing microcystin concentrations significantly delayed maturation, reduced size at first reproduction, number of eggs, and newborns. The interaction between carbaryl and Microcystis was highly significant. Animals matured later and at a smaller size than in controls. The number of eggs per female was reduced as well. Moreover, combined stressors caused frequent premature delivery of offspring with body deformations such as dented carapax or an undeveloped heart. This effect is concluded to be synergistic and could not be predicted from the effects of the single stressors.     

The impact of food quantity on UV-B tolerance and recovery from UV-B damage in Daphnia pulex

UV-B (290 nm) tolerance of Daphnia pulex, conditioned to four different food levels (Chlorophyta), was tested under standardized conditions with an artificial radiation source. Parameters measured were survival, percentage of egg bearing Daphnia and the number of juveniles produced after irradiation. UV-B tolerance of Daphnia pulex was found to be significantly improved with increasing food concentrations at all three parameters. The impact of the four different food concentrations on the photoreactivation system was tested with simultanous UV-B and white-light irradiation of Daphnia. Survival rate improved significantly with increasing food levels compared to solely UV-B irradiation. Photoreactivation had no effect on the reproductive parameters.     

Daphnia performance on diets containing different combinations of high‐quality algae,heterotrophic bacteria,and allochthonous particulate organic matter

1. Filter‐feeding zooplankton in lakes feed on a mixture of phytoplankton, bacteria, and terrestrial particles and the proportions and nutritional value of these components can be highly variable. However, the extent to which food quality interacts with food quantity in affecting overall zooplankton performance is not yet fully resolved.
2. Here we performed laboratory feeding experiments to test how the performance of the unselective filter feeder Daphnia galeata was affected if various quantities of high‐quality food (the phytoplankton Rhodomonas) were diluted with low‐quality food such as heterotrophic bacteria (Pseudomonas) or terrestrial detritus particles (t‐POM) from the riparian zone of a boreal forest stream. We hypothesised: that increased proportions of bacteria and t‐POM in the diet will lead to decreased survival, somatic growth; and reproduction of Daphnia despite the presence of phytoplankton; that these effects are more pronounced for t‐POM than for heterotrophic bacteria; and that this response is stronger when phytoplankton availability is low.
3. Increasing the concentrations of Pseudomonas affected Daphnia survival, growth, and reproduction negatively when Rhodomonas was available at intermediate (0.37 mgC/L) and high (0.55 mgC/L) quantities. When Rhodomonas quantity was low (0.22 mgC/L), the addition of Pseudomonas generally resulted in better Daphnia performance except at very high concentrations of the bacterium relative to Rhodomonas. In contrast, the addition of t‐POM was detrimental for overall Daphnia performance at all Rhodomonas concentrations.
4. Daphnia performance was best described by a model including the interaction between food quality and quantity, with stronger negative effects on Daphnia when high‐quality food was supplemented with t‐POM than with Pseudomonas.
5. The results indicate that the ability of zooplankton to use low‐quality food is affected by the concurrent availability of high‐quality food. Furthermore, food sources that can be used but do not fulfil dietary requirements of grazers (e.g. bacteria), may still provide nutritional benefits as long as other complementary food components are available in sufficient quantities to compensate for biochemical deficiencies.
6. Therefore, we conclude that heterotrophic bacteria, but not peat layer t‐POM, can be an important component of zooplankton diets in boreal lakes, especially if the concentration of phytoplankton is low.

     

Feeding at different plankton densities alters invasive bighead carp (<Emphasis Type="Italic">Hypophthalmichthys nobilis</Emphasis>) growth and zooplankton species composition

Invasive Asian carps Hypophthalmichthys spp. are an ecological threat to non-native aquatic ecosystems throughout the world, and are poised to enter the Laurentian Great Lakes. Little is known about how these filter-feeding planktivores grow and impact zooplankton communities in mesotrophic to oligotrophic systems like the Great Lakes. Our purpose was to determine how different plankton densities affect bighead carp H. nobilis biomass and how bighead carp affect zooplankton species composition. We conducted a 37-day indoor mesocosm experiment (volume = 678 l) with high and low plankton treatments (zooplankton dry mass ≈ 1,900 and 700 μg l⁻¹; chlorophyll a = 25 and 14 μg l⁻¹, respectively) in the presence and absence of juvenile bighead carp (mean = 5.0 g, 8.5 cm). Carp lost weight in the low plankton treatment and gained weight in the high plankton treatment, suggesting that food availability may be a limiting factor to bighead carp growth in regions of low plankton densities. In the presence of carp, zooplankton shifted from Daphnia to copepod dominance, while in the absence of carp, Daphnia remained dominant. Chydorids and ostracods increased in the presence of carp, but only in the low plankton treatment, suggesting that the impact of bighead carp on zooplankton species composition may vary with zooplankton density. Chlorophyll was higher in the absence of carp than in the presence. Chlorophyll and zooplankton densities in many Great Lakes ecosystems are substantially lower than our low treatment conditions, and thus our results suggest that Asian carp establishment in these regions may be unlikely. Handling editor: S. Declerck     

Genetic transformation and overexpression of a rice <Emphasis Type="Italic">Hd3a</Emphasis> induces early flowering in <Emphasis Type="Italic">Saussurea involucrata</Emphasis> Kar. et Kir. ex Maxim

Saussurea involucrata is a valuable traditional Chinese medicinal herb. This is the first report of a successful genetic transformation protocol for S. involucrata using Agrobacterium tumefaciens. Leaf explants were incubated with A. tumefaciens strain EHA105 harboring the binary vector pCAMBIA 1301, which contains the hpt gene as a selectable marker for hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, about 23.7% of the explants produced hygromycin-resistant calli on MS basal medium (Murashige and Skoog in Physiol Plant 15: 473–497, 1962) supplemented with 1 mg l⁻¹ benzyladenine (BA), 0.1 mg l⁻¹ α-naphthaleneacetic acid (NAA), 0.1 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D), 20 mg l⁻¹ hygromycin, and 500 mg l⁻¹ cefotaxime. Shoots were regenerated following transfer of the resistant calli to shoot induction medium containing 1.5 mg l⁻¹ BA, 0.1 mg l⁻¹ NAA, 0.25 mg l⁻¹ gibberellic acid (GA3), 20 mg l⁻¹ hygromycin, and 250 mg l⁻¹ cefotaxime, and about 67.5% of the resistant calli differentiated into shoots. Finally, 80% of the hygromycin-resistant shoots rooted on MS media supplemented with 0.2 mg l⁻¹ NAA, 20 mg l⁻¹ hygromycin, and 250 mg l⁻¹ cefotaxime. The transgenic nature of the transformants was demonstrated by detection of β-glucuronidase activity in the primary transformants and by Southern blot hybridization analysis. About 16% of the total inoculated leaf explants produced transgenic plants after approximately 5 months. Using this optimized transformation system, a rice ortholog of the Arabidopsis FLOWERING LOCUS T gene, Hd3a, was transferred into S. involucrata. Introduction of this gene caused an early-flowering phenotype in S. involucrata.     

Implications of phytoplankton chemical composition for zooplankton production: experimental evidence

Summary Growth and reproduction of twoDaphnia species (Cladocera) from Lake Constance were measured experimentally in a through-flow laboratory system under different food conditions, in order to investigate the effect of food quality on zooplankton populations. The cryptomonadRhodomonas sp. and the diatomStephanodiscus hantzschii were used as food. BothDaphnia species grew well at concentrations of 0.2–2.0 mgC/l ofRhodomonas sp., and 0.4–2.0 mgC/l ofS. hantzschii, whileStephanodiscus concentrations of 0.2 mgC/l did not supportDaphnia growth. Nitrogen content, which was very different in the two algae (C/N ratio = 4.9 forRhodomonas and 10.9 forS. hantzschii), was presumed to be the limiting factor. To test this hypothesis, nitrogen-limitedRhodomonas was offered to the cladocerans and their growth observed: the reduction in algal nitrogen content hindered animals' growth and reproduction. Results suggested that zooplankton production may also be affected by the chemical composition of the food algae, and that nitrogen potentially limits production, even in freshwater environments that are generally not N-limited.