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.

In vitro approaches to evaluate toxicity induced by organotin compounds tributyltin (TBT), dibutyltin (DBT), and monobutyltin (MBT) in neuroblastoma cells.

The toxic effects of the organotin compounds (OTCs) monobutyltin (MBT), dibutyltin (DBT), and tributyltin (TBT) were evaluated in vitro in a neuroblastoma human cell line. Mechanisms of cell death, apoptosis versus necrosis, were studied by using several markers: inhibition of cell viability and proliferation, F-actin, and mitochondrial membrane potential changes as well as reactive oxygen species (ROS) production and DNA fragmentation. The most toxic effects were detected with DBT and TBT even at very low concentrations (0.1-1 µM). In contrast, MBT induced lighter cytotoxic changes at the higher doses tested. None of the studied compounds stimulated propidium iodide uptake, although the most toxic chemical, TBT, caused lactate dehydrogenase release at the higher concentrations tested. These findings suggest that in neuroblastoma, OTC-induced cytotoxicity involves different pathways depending on the compound, concentration, and incubation time. A screening method for DBT and TBT quantification based on cell viability loss was developed, allowing a fast detection alternative to complex methodology.

Polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) and dioxin-like polychlorinated biphenyls (dl-PCBs) in fish, seafood products and fish oil in Spain.

A total of 84 samples of wild and farmed fish, cephalopods and fish oils for animal feeding, traded in Spain, were analysed for polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) and dioxin-like polychlorinated biphenyls (dl-PCBs) in 2009-2012, by gas chromatography-ion trap tandem mass spectrometry (GC-MS-MS). The method was optimised for screening at moderate costs, allowing PCDD/Fs determination at 1 pg World Health Organization-toxic equivalent quantities (WHO-TEQ) g⁻¹ wet weight (w w) and dl-PCBs at 0.02 pg WHO-TEQ g⁻¹ w w. Concentrations in fish and cephalopods ranged from values below the limit of detection to 1.7 pg g⁻¹ WHO-TEQ sum PCDD/Fs and dl-PCBs, considered as safe with regard to EU legislation. Higher levels were found in cod livers (5.4-54.2) and fish oils (3.3-30.7), with one noncompliant sample in each group.

Evaluation of an instrument for screening patients at risk for chronic kidney disease: testing SCORED (Screening for Occult Renal Disease) in a Portuguese population.

Prevalence of chronic kidney disease (CKD) is increasing and CKD has a long asymptomatic phase suitable for screening. SCORED (Screening for Occult Renal Disease) is a prescreening test which has compared favorably with KEEP. We report the results of SCORED testing in subjects attending a World Kidney Day event. After SCORED, subjects were tested for creatinine, urinary albumin and creatinine, and renal ultrasound. Eighty-eight subjects participated (32 men; mean age 59.7 ± 14.8 years; 58% hypertensive and 15.9% diabetics) of which 60 had a high score for kidney disease. Thirty-eight of 47 (80.8%) subjects that were further evaluated had a high-risk score. All subjects with CKD had a high score (100% sensitivity). SCORED showed low specificity (24.3%), but a high negative predictive value (100%). Including albuminuria in the definition of CKD increased the positive predictive value to 43.6%. In conclusion, SCORED is good for prescreening subjects for CKD in a European population as it captures all patients with CKD. Moreover, in subjects with low risk, the probability of CKD is low. SCORED is useful in alerting the general population and the medical community about the risk factors of CKD.

Cytotoxic activity of extracts of marine sponges from NW Spain on a neuroblastoma cell line.

Six species of marine sponges collected at intertidal and sublittoral sites of the coast of Galicia (NW Spain) were screened for potential cytotoxic properties on Neuroblastoma BE(2)-M17 cell line. Exposure to Halichondria panicea, Pachymatisma johnstonia, Ophlitaspongia seriata and Haliclona sp. aqueous extracts strongly affected cell appearance, inducing loss of neuron-like morphology and the formation of clumps. Extracts from these species also caused significant rates of cell detachment and decrease of mitochondrial membrane potential. Incubation with P. johnstonia, O. seriata and Suberites massa extracts also decreased the rate of cell proliferation. The increase of incubation time enhanced propidium iodide uptake by neuroblastoma cells. Toxic responses triggered by sponge extracts are compatible with apoptotic phenomena in neuroblastoma cells, even though increasing propidium uptake at long periods of exposure might indicate the induction of secondary necrosis. The cytotoxic properties of the tested extracts suggest the presence of compounds with potential pharmacological or biotechnological applications in the screened sponge species.

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.

Comparison between a TaqMan polymerase chain reaction assay and a culture method for ctx-positive Vibrio cholerae detection.

The main objective of the present work was to evaluate a real-time polymerase chain reaction (PCR) method to detect toxigenic Vibrio cholerae in Pangasius hypophthalmus, a freshwater fish cultured mainly in South East Asia. A FDA traditional culture method and a real-time PCR method of the ctx gene were used for detection of V. cholerae in spiked samples of pangasius fish. After an overnight enrichment of samples at 37 degrees C in alkaline peptone water, 2 cfu/25 g of fish was detected with both methods. Although both methods were very sensitive, obtaining results with culture methods may take several days, while real-time PCR takes only a few hours. Furthermore, with traditional methods, complementary techniques such as serotyping, although not available for all serogroups, are needed to identify toxigenic V. cholerae. However, with real-time PCR, toxigenic serogroups are detected in only one step after overnight enrichment.

Enriching rotifers with "premium" microalgae. Nannochloropsis gaditana.

The nutritive quality of Nannochloropsis gaditana cultured semicontinuously with different daily renewal rates was tested as a diet for short-term enrichment of the rotifer Brachionus plicatilis. After 24 h, dramatic differences in the survival, dry weight, and biochemical composition of the rotifers depending on the renewal rate of microalgal cultures were observed. Survival after the feeding period increased with increasing renewal rates. Rotifers fed microalgae from low renewal rate, nutrient-deficient cultures showed low dry weight and organic contents very similar to those of the initial rotifers that were starved for 12 h before the start of the feeding period. On the contrary, rotifers fed nutrient-sufficient microalgal cells underwent up to twofold increases of dry weight and protein, lipid, and carbohydrate contents with regard to rotifers fed nutrient-depleted N. gaditana. Consequently, feed conversion rate decreased in these conditions, indicating a better assimilation of the microalgal biomass obtained at high renewal rates. No single microalgal biochemical parameter among those studied can explain the response of the filter feeder. Similarly to gross composition, EPA and n-3 contents in rotifers fed microalgae from nutrient-sufficient cultures were double than the contents found in rotifers fed nutrient-limited microalgae. In addition, very high positive correlations between the contents of EPA and n-3 in N. gaditana and B. plicatilis were observed. These results demonstrate that selecting the appropriate conditions of semicontinuous culture can strongly enhance the nutritional value of microalgae that is reflected in the growth and biochemical composition of the filter-feeder even in short exposure periods.