Chemical composition of the sponge Chondrosia reniformis from [2pt] the Canary Islands

Lake Zempoala was studied throughout 16 months in 1996–1997. It is a shallow monomictic lake situated at 2800 masl at the Neovolcanic Belt, well within the Mexican tropical zone. Most of the phytoplankton species in this lake may be characterized as temperate, according to their geographical distribution. A break down in phytoplankton biomass was observed before the lake's circulation, and open to question if a clear-water phase could be present in a tropical lake.     

Polyamines in marine macroalgae: Levels of putrescine, spermidine and spermine in the thalli and changes in their concentration during glycerol-induced cell growth in vitro

The levels of putrescine (Put), spermidine (Spd) and spermine (Spm) were analyzed in naturally collected samples of the marine macroalgae Dyctiota dichotoma, Gelidium canariensis and Grateloupia doryphora . Polyamines (PAs) appeared in free (35–134 μg g⁻¹ fresh weight) and bound TCA-insoluble form (1 667–2 624 μg g⁻¹ fresh weight). Axenic in vitro cultures of sporelings from G. doryphora were established in the medium containing glycerol. This medium promoted growth and morphogenesis and also increased the free and bound PA levels in the sporelings. Tracer experiments using 70 kBq [U-¹⁴C]-glycerol showed significant quantities of radioactivity in Put, Spd and Spm after 20 h of incubation. The effects of glycerol on growth were inhibited by the ornithine decarboxylase (EC 4.1.1.17) inhibitor α -difluoromethylornithine (DFMO). The presence of DFMO in the incubation medium with [U-¹⁴C]-glycerol also reduced the radioactivity in PAs.     

Analysis of ethylene‐induced gene regulation during carposporogenesis in the red seaweed Grateloupia imbricata (Rhodophyta)

Ethylene favors carposporogenesis in the red seaweed Grateloupia imbricata. Analyses of cystocarp development in vitro in thalli treated with ethylene suggest an interconnection between polyamine and ethylene biosynthesis pathways. Yet, little is known about molecular mechanisms underlying carposporogenesis. Here, we used droplet digital PCR to analyze genes encoding enzymes related to polyamine (Spermidine [Spd] synthase) and ethylene (ACC synthase) synthesis; a pivotal compound of both pathways (S‐adenosyl methionine synthase, SAMS); the gene that encodes amine oxidase, which is involved in polyamine degradation, and a candidate gene involved in seaweed reproduction (ornithine decarboxylase, ODC). In addition, we analyzed genes encoding proteins related to stress and reactive oxygen species, ascorbate peroxidase (APX), cytochrome P450 and WD 40. We characterized gene expression in fertilized and fertile thalli from G. imbricata that were exposed to ethylene for 15 min at two time points after treatment (1 and 7 d). The differential gene expression of SAMS, Spd synthase, ACC synthase, and cytochrome P450 was related to disclosure and development of cystocarps in fertilized thalli that transitioned from having no visible cystocarps at 1 d to developing cystocarps at 7 d. Likewise, cytochrome P450 was associated with cystocarp disclosure and maturation. In addition, amine oxidase and APX were involved in fine‐tuning polyamine and reactive oxygen species during carposporogenesis, respectively, whereas WD 40 did so in relation to ethylene signaling. Expression of the candidate gene ODC was increased when cystocarps were not visible (fertilized thalli, 1d), as previously described. This analysis suggests developmental stage‐specific roles for these genes during carposporogenesis.     

Influence of plant growth regulators, polyamines and glycerol interaction on growth and morphogenesis of carposporelings of Grateloupia cultured in vitro

The influence of the plant growth regulators 2,4-D, GA₃, BA and kinetin, and the polyamines putrescine, spermidine and spermine were tested on axenic in vitro cultures of carposporelings of Grateloupia doryphora. The auxin 2,4-D (10^-3 M) and the polyamine spermine (10^-6 M and 10^-3 M) induced a callus (disorganised cell mass that arose from the organised tissue of the carposporeling, as demonstrated by microscopic monitoring of the tissue). Putrescine and spermidine (10^-3 M) transformed the carposporelings into cell masses that produced shoots. BA (10^-3 M) and kinetin (10^-6 M and 10^-3 M) were inhibitory. In 10^-1 M glycerol-containing culture medium, which is known to induce the formation of morphogenic cell masses, the addition of GA₃ M) resulted in the inhibition of the morphogenesis (i.e. shoot emission) in the cell mass. The kinetin at 10^-6 M inhibited morphogenesis, whilst at 10^-3 M inhibited even the formation of the cell masses. The combination of glycerol (10^-1 M) and the auxin 2,4-D (10^-6 and 10^-3 M) or the polyamines putrescine, spermidine and spermine (10^-6 and 10^-3 M) resulted in a bigger size of the cell masses that led to a higher amount of shoots per cell mass than in glycerol alone. This revised version was published online in June 2006 with corrections to the Cover Date.     

POLYAMINES INFLUENCE MATURATION IN REPRODUCTIVE STRUCTURES OF GRACILARIA CORNEA (GRACILARIALES,RHODOPHYTA)1

Naturally occurring levels of putrescine, spermidine, and spermine were analyzed in female gametophyte (nonfertilized) and reproductive tissues (cystocarps) at two different stages of development in the marine red algae Gracilaria cornea J. Agardh. Endogenous polyamine levels changed at differential stages of cystocarp maturation. Highest polyamine values were found on tissue from the early post‐fertilization stage, decreasing as the cystocarp matured. Incubation experiments revealed that exogenous polyamines induced cystocarp maturation and promoted carpospore liberation, developing cell masses within 4 to 7 days in treatments with spermine. This is the first report on the effect of polyamines on cystocarp maturation in marine algae.     

Effect of dietary lipids on plasma fatty acid profiles and prostaglandin and leptin production in gilthead seabream (Sparus aurata)

The aim of this study was to investigate the effects of different levels of substitution of fish oil by vegetable oils rich in oleic, linoleic and linolenic acids on gilthead seabream plasma and leukocyte fatty acid compositions and prostaglandin (PG) and leptin production. Juvenile seabream of 24 g initial body mass were fed four iso-energetic and iso-proteic experimental diets for 281 days. Fatty acid composition of plasma lipids was markedly affected by the inclusion of vegetable oils (VO). ARA (arachidonate), EPA (eicosapentaenoate) and DHA (docosahexaenoate) were preferentially incorporated into polar lipids of plasma, and DHGLA (di-homogammalinoleate) accumulated with increased vegetable oil inclusion. Dietary treatments resulted in alterations of DHGLA/ARA ratios, but not ARA/EPA. ARA-derived PGE₂ production in plasma was not affected by vegetable oils, in agreement with similar eicosanoid precursor ratio (ARA/EPA) in leukocytes total lipids and plasma phospholipids among fish fed with the different dietary treatments. Feeding vegetable oils leads to a decrease in plasma EPA which in turn reduced plasma PGE₃ concentration. Moreover, PGE₃ was the major prostaglandin produced in plasma of fish fed fish oil based diet. Such findings point out the importance of EPA as a precursor of prostaglandins in marine fish, at least for the correct function of the blood cells, and correlates well with the predominant role of this fatty acid in immune regulation in this species. A negative correlation was found between plasma PGE₂ and leptin plasma concentration, suggesting that circulating levels of leptin may act as a metabolic signal modulating PGE₂ release. The present study has shown that increased inclusion of vegetable oils in diet for gilthead seabream may profoundly affect the fatty acid composition of plasma and leukocytes, specially HUFA (highly unsaturated fatty acids), and consequently the production of PGE₃, which can be a major PG in plasma. Alteration in the amount and type of PG produced can be at least partially responsible for the changes in the immune system and health parameters of fish fed diets with high inclusion of VO.     

Effect of Spirostane Analogues of Brassinosteroids on Callus Formation and Plant Regeneration in Lettuce (Lactuca sativa)

Two spirostane analogues of brassinosteroids (BB-6 and MH-5) were tested for callus induction and plant regeneration in lettuce. They were used as a cytokinin (6-BA) substitute or in combination with 6-BA at different concentrations. Treatment with 0.1 mg l⁻¹ 6-BA was used as control. Results showed that there was no callus induction when 6-BA was substituted by these analogues. However, BB-6 and MH-5 enhanced both callus formation and shoot regeneration from cotyledons in lettuce when added at determined concentrations with 0.1 mg l⁻¹ of 6-BA in the culture medium.     

Sporulation and sterilization method for axenic culture of Gelidium canariensis

A sporulation and sterilization procedure was used to establish axenic cultures of sporelings of Gelidium canariensis. Sporangial branchlets excised from the thallus were rinsed in distilled water twice and in 1% sodium hypochlorite (2 min). The branchlets were cultivated overnight in multiwell plates with 0.3 ml of autoclaved seawater to promote spore liberation in 90% of the cultivated branchlets. The branchlets were transferred to an antibiotic solution made of ampicillin, penicillin, rifampicin, nystatin (0.2 mg ml⁻¹ each) and 0.1 g ml⁻¹ of GeO₂ in liquid PES for 45 days, during which clusters of spores (85–100 spores) were observed on the surface of the branchlet. After 55 days, they became axenic sporelings with the prostrate and erect system characteristic of Gelidium canariensis.     

Molecular mechanisms underlying Grateloupia imbricata (Rhodophyta) carposporogenesis induced by methyl jasmonate

When applied in vitro, methyl jasmonate is sensed by the red seaweed Grateloupia imbricate, substantially and visually affecting its carposporogenesis. However, although there is some understanding of the morphological changes induced by methyl jasmonate in vitro, little is known about the genes that are involved in red seaweed carposporogenesis and how their protein products act. For the work reported herein, the expression of genes in red seaweed that encode enzymes involved in the synthesis of methyl jasmonate (jasmonic acid carboxyl methyl transferase and a putative methyl transferase) was monitored. Additionally the genes involved in oxidation (cytochrome P450 and WD40), jasmonate synthesis, signal transduction, and regulation of reactive oxygen species (MYB), and reproduction (ornithine decarboxylase) were monitored. To determine when or if the aforementioned genes were expressed during cystocarp development, fertilized and fertile thalli were exposed to methyl jasmonate and gene expression was measured after 24 and 48 h. The results showed that methyl jasmonate promoted differential gene expression in fertilized thalli by 24 h and upregulated expression of the ornithine decarboxylase gene only by 48 h in fertile thalli (0.75 ± 003 copies · μL^?1 at 24 h vs. 1.11 ± 0.04 copies · μL^?1 at 48 h). We conclude that Ornithine decarboxylase expression involves methyl jasmonate signaling as well as development and maturation of cystocarps.     

Morphogenetic effect of glycerol on tissue cultures of the red seaweed Grateloupia doryphora

Explants of Grateloupia doryphora were cultivated in Provasoli Enriched Seawater culture medium (PES) supplemented with glycerol (0.1, 0.3, 0.5 or 0.8 mol 1^–1) or carbohydrates (0.1 or 0.3 mol 1^–1 mannose, glucose and galactose) and agar (3, 8, 15 g 1^–1 ). The osmolality of the medium was adjusted by dilution of the seawater (70 or 100%, v/v). The increase in fresh weight of explants cultivated in liquid medium with glycerol (0.3 mol 1^–1) and without glycerol was compared. All experiments were carried out in the light, except for one assay in which the explants were cultivated in the dark. Glycerol was an effective carbon source for the vegetative propagation of G. doryphora in solid and liquid media. Mannose, glucose and galactose all had no effect on growth or morphogenesis of the explants. In solid media the main effect of glycerol was as a morphogenetic inductor, with PES70 (70% seawater) + 0.1 or 0.3 mol 1^–1 glycerol + 3 or 8 g 1^–1 agar the best formulation. An increase in the concentration of agar in glycerol-containing medium reduced the morphogenetic capacity of the explants, which developed into compact cell masses. The effects of glycerol were observed only in explants cultivated under light.