The potential of fatty acid isotopes to trace trophic transfer in aquatic food-webs.

Compound-specific isotope analyses (CSIA) of fatty acids (FA) constitute a promising tool for tracing energy flows in food-webs. However, past applications of FA-specific carbon isotope analyses have been restricted to a relatively coarse food-source separation and mainly quantified dietary contributions from different habitats. Our aim was to evaluate the potential of FA-CSIA to provide high-resolution data on within-system energy flows using algae and zooplankton as model organisms. First, we investigated the power of FA-CSIA to distinguish among four different algae groups, namely cyanobacteria, chlorophytes, haptophytes and diatoms. We found substantial within-group variation but also demonstrated that δ13C of several FA (e.g. 18:3ω3 or 18:4ω3) differed among taxa, resulting in group-specific isotopic fingerprints. Second, we assessed changes in FA isotope ratios with trophic transfer. Isotope fractionation was highly variable in daphnids and rotifers exposed to different food sources. Only δ13C of nutritionally valuable poly-unsaturated FA remained relatively constant, highlighting their potential as dietary tracers. The variability in fractionation was partly driven by the identity of food sources. Such systematic effects likely reflect the impact of dietary quality on consumers' metabolism and suggest that FA isotopes could be useful nutritional indicators in the field. Overall, our results reveal that the variability of FA isotope ratios provides a substantial challenge, but that FA-CSIA nevertheless have several promising applications in food-web ecology. This article is part of the theme issue 'The next horizons for lipids as 'trophic biomarkers': evidence and significance of consumer modification of dietary fatty acids'.

Enriching Rotifers with "Premium" Microalgae: Rhodomonas lens.

The nutritional value of the marine cryptophyte Rhodomonas lens for the filter feeder Brachionus plicatilis as well as its biotechnological potential as a source of phycoerythrin (PE) and polyunsaturated fatty acids (PUFA) were evaluated in semi-continuous cultures maintained with different daily renewal rates (RR), from 10% (R10) to 50% (R50) of the total volume. Steady-state cell density decreased from 22 to 7 × 106 cells mL-1 with increasing RR, with the maximum cell productivity, nearly 0.4 g L-1 day-1, observed with R40. PE cell content attained the highest values with the highest RR (circa 9 pg cell-1). All treatments of R. lens maintained under nitrate-saturated conditions (R20-R50) showed a similar high content of PUFAs, > 60% of total fatty acids (FA), with linolenic acid (18:3n-3) and 18:4n-3, representing 12 and 29% of total FA respectively. The PUFA level in the nitrogen-limited R10 cultures was significantly lower (37%). R. lens promoted higher weight gain in the rotifer B. plicatilis than Tisochrysis lutea (T-ISO), a species commonly used for rotifer culture and enrichment. Significant differences were found in the protein content and in the ratio n-3/n-6 fatty acids among rotifers fed with R. lens from different RRs, with higher values being found in those fed with R. lens from higher RRs. The enrichment of the rotifers for short periods of 3 h was sufficient to modify the biochemical composition of the rotifers, but it was evidenced as too short for the accumulation of PUFAs, when compared to long-term (24 h) enrichment. The rotifers reflected the higher protein and PUFA content of R. lens cultivated with nutrient sufficient microalgae (R40) after only 3 h of enrichment. These results demonstrate that semi-continuous culture of R. lens under appropriate conditions can strongly enhance the nutritional value of this species, being reflected in the growth and biochemical composition of the filter feeder, even in short exposure periods.

A rotifer-derived paralytic compound prevents transmission of schistosomiasis to a mammalian host.

Schistosomes are parasitic flatworms that infect over 200 million people, causing the neglected tropical disease, schistosomiasis. A single drug, praziquantel, is used to treat schistosome infection. Limitations in mass drug administration programs and the emergence of schistosomiasis in nontropical areas indicate the need for new strategies to prevent infection. It has been known for several decades that rotifers colonizing the schistosome's snail intermediate host produce a water-soluble factor that paralyzes cercariae, the life cycle stage infecting humans. In spite of its potential for preventing infection, the nature of this factor has remained obscure. Here, we report the purification and chemical characterization of Schistosome Paralysis Factor (SPF), a novel tetracyclic alkaloid produced by the rotifer Rotaria rotatoria. We show that this compound paralyzes schistosome cercariae and prevents infection and does so more effectively than analogous compounds. This molecule provides new directions for understanding cercariae motility and new strategies for preventing schistosome infection.

The ecological competition and grazing reverse the effects of sulfamethoxazole on plankton: a case study on characterizing community-level effect.

The toxic effects of sulfamethoxazole (SMX) on densities of two algae, Platymonas helgolandica var. tsingtaoensis, Isochrysis galbana, and of a rotifer, Brachionus plicatilis, were tested by the population and community experiments. Two endpoints, the carrying capacity and the densities array of community in steady state (DACS), were used to characterize the toxic effects at a population level and a community one, respectively. The results showed that the carrying capacity of P. helgolandica var. tsingtaoensis and B. plicatilis did not decline significantly in population test when the concentration of SMX was lower than 6.0 mg L-1 and 12.0 mg L-1, respectively. However, I. galbana was sensitive to SMX and had presented toxic effect at 3.0 mg L-1. By extrapolation of toxic effect at a population level to a community one, a derived community-NOEC was 3.0 mg L-1, representing an inference from data of toxic effects at population level. In community experiment, when the customized community was in steady state, the density of I. galbana increased as a whole with SMX concentration in the range of tested concentration (0-144 mg L-1), while that of P. helgolandica var. tsingtaoensis assumed the trend of a reversed "v" in this range. Only the density of B. plicatilis decreased with SMX concentration. With the DACS as endpoint, a NOEC for the customized community was determined to be 6.0 mg L-1. This indicates that interspecific interactions can reverse the toxic effects of SMX on phytoplankton. The DACS was reliable and stable, serving as the endpoints in assessment of the effects of the pollutants on the ecosystems.

Aging extension and modifications of lipid metabolism in the monogonont rotifer Brachionus koreanus under chronic caloric restriction.

To examine the interrelationship of aging extension and modification of lipid metabolism under chronic caloric restriction (CCR; reduced concentration of the green algae Tetraselmis suecica) in the monogonont rotifer Brachionus koreanus, we assessed life cycle parameters, fatty acid composition, and expression of sirtuin and genes related to lipid metabolism. B. koreanus in the 5% T. suecica group showed an increased life span but decreased reproduction. Based on this finding, we chose 5% T. suecica for further experiments and compared the data with those for 100% T. suecica. Upregulation of sirtuin gene expression was observed under CCR. In addition, despite the reduction in the amount of total fatty acid (FA) and the area of triacylglycerol, increases in the ratios of saturated fatty acid and monounsaturated fatty acid (MUFA) to total FA in 5%-exposed B. koreanus were observed. Furthermore, mRNA expression analysis confirmed that CCR promoted the synthesis of MUFA through Δ9 desaturase. Moreover, expression of the docosahexaenoic acid (DHA) synthesizing gene Δ4 desaturase was also upregulated, together with DHA content. These data suggest that CCR modified protein acetylation and lipid metabolism, leading to a decrease in reproduction and consequently resulting in life span extension.

The derivation of effects threshold concentrations of lead for European freshwater ecosystems.

The main objective of the present study was to derive ecologically relevant effect threshold concentrations of (dissolved) Pb for selected European Union (EU) freshwater rivers, using the 2008 EU Voluntary Risk Assessment Report as a starting point and more advanced methodologies than those used in the Voluntary Risk Assessment Report. This included 1) implementing more robust quality criteria for selecting chronic toxicity data; 2) the conversion of total to dissolved Pb concentrations using a combination of an empirical equation relating inorganic Pb solubility and geochemical speciation modeling to account for effects of dissolved organic matter; 3) the use of bioavailability models for chronic toxicity for species belonging to 3 different trophic levels; and 4) the use of robust methods for large data set handling (such as species sensitivity distribution [SSD] analysis). The authors used published bioavailability models for an algal species (Pseudokirchneriella subcapitata) and a daphnid (Ceriodaphnia dubia) and developed a new model for the fathead minnow (Pimephales promelas). The research has shown that these models are also useful for, and reasonably accurate in, predicting chronic toxicity to other species, including a snail, a rotifer, midge larvae, and an aquatic plant (read-across). A comprehensive chronic toxicity data set for Pb was compiled, comprising 159 individual high-quality toxicity data for 25 different species. By applying the total dissolved conversion and the bioavailability models, normalized toxicity values were obtained, which were then entered into a SSD analysis. Based on the parametric best-fitting SSDs, the authors calculated that ecological threshold concentrations of Pb protecting 95% of freshwater species for 7 selected European freshwater scenarios were between 6.3 µg dissolved Pb/L and 31.1 µg dissolved Pb/L.

Impact of arachidonic acid enrichment of live rotifer prey on bacterial communities in rotifer and larval fish cultures.

Rotifers (Brachionus plicatilis), commonly used at first feeding in commercial fish hatcheries, carry a large bacteria load. Because they are relatively poor in essential fatty acids, it is common practice to enrich them with fatty acids, including arachidonic acid (AA). This study aims to determine whether prey enrichment with AA may act as a prebiotic and modify the microbial community composition either in AA-enriched rotifer cultures or in larval-rearing water using winter flounder (Pseudopleuronectes americanus) as a larval fish model. AA enrichment modified the bacterial community composition in both the rotifer culture tanks and the larval-rearing tanks. We observed an increase in the number of cultivable bacteria on TCBS (thiosulfate-citrate-bile salts-sucrose) agar, used as a proxy for the abundance of Vibrio sp. The results suggest that AA may also play an indirect role in larval health.

Anions mediate ligand binding in Adineta vaga glutamate receptor ion channels.

AvGluR1, a glutamate receptor ion channel from the primitive eukaryote Adineta vaga, is activated by alanine, cysteine, methionine, and phenylalanine, which produce lectin-sensitive desensitizing responses like those to glutamate, aspartate, and serine. AvGluR1 LBD crystal structures reveal an unusual scheme for binding dissimilar ligands that may be utilized by distantly related odorant/chemosensory receptors. Arginine residues in domain 2 coordinate the γ-carboxyl group of glutamate, whereas in the alanine, methionine, and serine complexes a chloride ion acts as a surrogate ligand, replacing the γ-carboxyl group. Removal of Cl(-) lowers affinity for these ligands but not for glutamate or aspartate nor for phenylalanine, which occludes the anion binding site and binds with low affinity. AvGluR1 LBD crystal structures and sedimentation analysis also provide insights into the evolutionary link between prokaryotic and eukaryotic iGluRs and reveal features unique to both classes, emphasizing the need for additional structure-based studies on iGluR-ligand interactions.

Nucleotide enrichment of live feed: a promising protocol for rearing of Atlantic cod Gadus morhua larvae.

We investigated the effect of two commercial nucleotide products (NT1 and NT2), administered through live feed, on growth and stress tolerance of Atlantic cod larvae. Expression of genes related to muscle growth (igf-1, igf1r, igf-2, fst, fgf6, myod, and myhc) and nucleotide metabolism (uox, hprt, ndk, and uck) was evaluated during larval development. In addition, the expression of genes related to stress (hif-1α, hif-2α, hif-3α, and mb) was studied after an air exposure stress test. The enrichment of rotifers with nucleotides did not reveal any difference in nucleotide profiles, the exception being the RNA level of the NT1-enriched group that was significantly higher than the unenriched rotifer. Unenriched Artemia showed poor nucleotide profiles compared to enriched Artemia since 5' UMP, 5' GMP, and 5' AMP were observed only in the nucleotide groups. At 38 days post-hatch (dph), NT1 group had significantly higher dry weight (3.1 ± 0.1 mg) than the control (CON; 2.3 ± 0.1 mg). The treatments did not produce any significant differences in the expression of the key myogenic genes. Among the genes associated with nucleotide metabolism, ndk was down-regulated in NT1 at 38 dph. In the air exposure test, survival was significantly higher in the CON (77 ± 6 %) than in NT1 (48 ± 3 %) and NT2 (50 ± 3 %). After air exposure, mb was expressed at lower levels in NT2 group, hif-2α was induced in NT1 group, and hif-3α was upregulated in all groups. Our findings indicate that the improvement in the nucleotide profile of Artemia upon nucleotide enrichment could eventuate in the rapid growth of larvae.

Thermostable proteins in the diapausing eggs of Brachionus manjavacas (Rotifera).

Diapausing embryos (resting eggs) from brachionid rotifers are able to withstand desiccation and thermal stress. Resting eggs can remain viable for decades, and develop normally once placed in a permissive environment that allows for hatching, growth and development. The exact mechanisms of resistance are not known, although several molecules have been suggested to confer protection during desiccation and thermal stress. In this study, we have identified by mass spectrometry two thermostable proteins, LEA (late embryogenesis abundant) and VTG (vitellogenin-like), found exclusively in the resting eggs of Brachionus manjavacas. This is the first observation that LEA proteins may play a role in thermostability and the first report of a VTG-like protein in the phylum Rotifera. These proteins exhibited increased expression in rotifer resting eggs when compared to amictic females. Our data suggest the existence of alternate pathways of desiccation and thermal resistance in brachionid rotifers.

Preservation, development, and hatching of resting eggs in the freshwater rotifer Brachionus calyciflorus pallas.

The resting eggs of Brachionus calyciflorus produced under laboratory by manipulating various environmental conditions were subjected to preservation for long-term storage. Preservation strategies included as storing freshly collected resting eggs, dry resting eggs, and storing at 4°C. Results showed that the resting eggs preserved by drying as they were able to maintain the viability without much variation in the hatchability. The scanning electron micrographs clearly depicted the surface damage as well as change in the shape and surface topography due to the storage. The resting eggs thus stored were traced for their normal embryonic development, during which as many as 11 stages were traced and time taken for each stage of development was observed. Microscopic observations revealed that the first four stages represented the development of embryo and thereafter around 16-18 h extrution of neonate from egg occurred. At 18-19 h of development the characteristic of anterior and posterior spines appeared in the neonate. Just released neonate, after 4-5 h of development reached adult stage. Overall the hatching and embryonic development of the resting eggs preserved by "drying" were normal and hence this method could be adopted for long-term storage.

Effect of the nutritional status of semi-continuous microalgal cultures on the productivity and biochemical composition of Brachionus plicatilis.

The rotifer Brachionus plicatilis was cultured using the microalga Isochrysis aff. galbana clone T-ISO as feed. T-ISO was cultured semi-continuously with daily renewal rates of 10%, 20%, 30%, 40%, and 50% of the volume of cultures. The increase of renewal rate led to increasing nutrient and light availability in microalgal cultures, which caused differences in the biochemical composition of microalgal biomass. Growth rate, individual dry weight, organic content, and biomass productivity of rotifer cultures increased in response to higher growth rate in T-ISO cultures. Rotifer growth rate showed a strong negative correlation (R² = 0.90) with the C/N ratio of microalgal biomass. Rotifer dry weight was also affected by nutrient availability of T-ISO cultures, increasing up to 50% from nutrient-limited to nutrient-sufficient conditions. Consequently, biomass productivity of rotifer cultures increased more than twofold with the increase of renewal rate of T-ISO cultures. Rotifer organic content underwent the same trend of total dry weight. Maximum content of polyunsaturated fatty acids was reached in rotifers fed T-ISO from the renewal rate of 40%, with percentages of docosahexaenoic acid (22:6ω-3, DHA) and eicosapentaenoic acid (20:5ω-3, EPA) of 11% and 5% of total fatty acids, respectively. Selecting the most appropriate conditions for microalgal culture can therefore enhance the nutritive quality of microalgal biomass, resulting in a better performance of filter feeders and their nutrient content, and may constitute a useful tool to improve the rearing of fish larvae and other aquaculture organisms that require live feed in some or all the stages of their life cycle.