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Mar Drugs
2021 Apr 17;194:. doi: 10.3390/md19040230.
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Two Benthic Diatoms, Nanofrustulum shiloi and Striatella unipunctata, Encapsulated in Alginate Beads, Influence the Reproductive Efficiency of Paracentrotus lividus by Modulating the Gene Expression.
Glaviano F
,
Ruocco N
,
Somma E
,
De Rosa G
,
Campani V
,
Ametrano P
,
Caramiello D
,
Costantini M
,
Zupo V
.
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Physiological effects of algal metabolites is a key step for the isolation of interesting bioactive compounds. Invertebrate grazers may be fed on live diatoms or dried, pelletized, and added to compound feeds. Any method may reveal some shortcomings, due to the leaking of wound-activated compounds in the water prior to ingestion. For this reason, encapsulation may represent an important step of bioassay-guided fractionation, because it may assure timely preservation of the active compounds. Here we test the effects of the inclusion in alginate (biocompatible and non-toxic delivery system) matrices to produce beads containing two benthic diatoms for sea urchin Paracentrotus lividus feeding. In particular, we compared the effects of a diatom whose influence on P. lividus was known (Nanofrustulum shiloi) and those of a diatom suspected to be harmful to marine invertebrates, because it is often present in blooms (Striatella unipunctata). Dried N. shiloi and S. unipunctata were offered for one month after encapsulation in alginate hydrogel beads and the larvae produced by sea urchins were checked for viability and malformations. The results indicated that N. shiloi, already known for its toxigenic effects on sea urchin larvae, fully conserved its activity after inclusion in alginate beads. On the whole, benthic diatoms affected the embryogenesis of P. lividus, altering the expression of several genes involved in stress response, development, skeletogenesis and detoxification processes. Interactomic analysis suggested that both diatoms activated a similar stress response pathway, through the up-regulation of hsp60, hsp70, NF-κB, 14-3-3 ε and MDR1 genes. This research also demonstrates that the inclusion in alginate beads may represent a feasible technique to isolate diatom-derived bioactive compounds.
Figure 1. Percentages of normal and malformed plutei derived from sea urchins fed with the control diet (U. rigida), N. shiloi and S. unipunctata encapsulated in alginate beads. One-way ANOVA followed by Tukey post hoc for multiple comparisons: **** p < 0.0001. Pairwise comparison was reported between control (U. rigida diets) vs. samples treated with N. shiloi and S. unipunctata.
Figure 2. Fold changes of stress response genes in plutei from sea urchin adults fed with N. shiloi (black bar) and S. unipunctata (gray bar) encapsulated in alginate beads. The dotted red line represents the cut-off (1.5). Values are reported as the average fold changes ± SD (n = 3). Statistical differences were evaluated by nonparametric Mann–Whitney test. p-values < 0.05 were considered significant.
Figure 3. Fold changes of development and differentiation genes in plutei from sea urchin adults fed with N. shiloi (black bar) and S. unipunctata (gray bar) encapsulated in alginate beads where (a) the two diatoms induced the same gene expression (up- or down-regulation for both diatoms) or (b) different gene expression (up- in one diatom and down-regulation in the other diatom). The dotted red line represents the cut-off (1.5). Values are reported as the average fold changes ± SD (n = 3). Statistical differences were evaluated by nonparametric Mann–Whitney test. p-values < 0.05 were considered significant.
Figure 4. Fold changes of skeletogenesis genes in plutei from sea urchin adults fed with N. shiloi (black bar) and S. unipunctata (gray bar) encapsulated in alginate beads. The dotted red line represents the cut-off (1.5). Values are reported as the average fold changes ± SD (n = 3). Statistical differences were evaluated by nonparametric Mann–Whitney test. p-values < 0.05 were considered significant.
Figure 5. Fold changes of detoxification genes in plutei from sea urchin adults fed with N. shiloi (black bar) and S. unipunctata (gray bar) encapsulated in alginate beads. The dotted red line represents the cut-off (1.5). Values are reported as the average fold changes ± SD (n = 3). Statistical differences were evaluated by nonparametric Mann–Whitney test. p-values < 0.05 were considered significant.
Figure 6. Gene network performed by STRING interactome of protein–protein interactions. Correlations with high confidence score cut-off (900) were reported. Among functionally correlated genes, those with up (red), down (green) and unchanged (blue) expression affected by N. shiloi (a) and S. unipunctata (b) were reported. Color shading depends on fold-change values. Gray spheres represent additional connections.
Figure 7. Gene network performed by STRING interactome of protein–protein interactions. Correlations with high confidence score cut-off (900) were reported. Among functionally correlated genes, those with up (red), down (green) and unchanged (blue) expression affected by N. shiloi (a) and S. unipunctata (b) were reported. Color shading depends on fold change values. Gray spheres represent additional connections.
Figure 8. Gene network performed by STRING interactome of protein–protein interactions. Correlations with high confidence score cut-off (900) were reported. Among functionally correlated genes, those with up (red), down (green) and unchanged (blue) expression affected by N. shiloi (a) and S. unipunctata (b) were reported. Full color shows greater fold change values. Gray spheres represent additional connections.
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