A. Aguilera

The use of polyclonal antisera and blocking of antibodies in the identification of marine dinoflagellates: species-specific and clone-specific antisera against Gymnodinium and Alexandrium

Abstract Separate polyclonal antibodies were developed against cell surface antigens of the 4 dinoflagellate species: (a) Gymnodinium catenatum ; (b) a morphologically similar non-toxic dinoflagellate Gyrodinium sp.; (c) Alexandrium minutum ; and (d) morphologically similar A. lusitanicum . Blocking of antibodies to obtain clone-specificity were performed. Positive immunofluorescence reactions were visualized with epifluorescence microscopy as well as spectrofluorometry using FITC indirect immunofluorescence assay. Specificity and cross reactivity were tested with cells from laboratory cultures as well as cells from natural samples. Polyclonal antisera were species-specific (for G. catenatum and Gyrodinium sp.) and genus-specific (for Alexandrium ). After blocking the antisera were clone-specific. An excellent labelling was obtained with fresh cells and from those preserved in 4% buffered formaldehyde. The blocking of the antibodies shows great promise as a technique to obtain clone-specific antisera.

Use of lectins to recognize and differentiate unicellular algae

Clonal cultures of five species of unicellular algae from three different phyla (Cyanobacteria, two Chroococcus spp.; Dinophyceae, two Prorocentrum lima; Conjugatophyceae, four Spirogyra insignis and two Zygnema spp.) were recognized using nine FITC labeled lectins: Phaseolus limensis (PHA), Phytolacca americana (PWM), Erithryna cristagalli (ECA), Canavalia ensiformis (Con A), Helix pomatia (HPA), Triticum vulgare (WGA), Pisum sativum (PEA), Arachis hypogaea (PNA) and Glycine maxima (SBA). The binding activity of lectins was easily detected by fluorescence, but some lectins were also able to agglutinate cells. Lectins appear to be an interesting tool to identify and differentiate between different clones (genotypes) of the same species. Also, lectins are able to discriminate between phylogenetically closely related species

The influence of the slowing of Earth's rotation: A hypothesis to explain cell division synchrony under different day duration in earlier and later evolved unicellular algae

Every year the Earths rotation period is reduced, mainly due to the tidal drag of the moon. The length of day increases continuously by about 1 h every 200 million years. The period of rotation around the Sun remains constant; hence, the length of the year remains constant, so years acquire progressively fewer days. Many unicellular algae show rhythmicity in their cell division cycle. If primitive algae evolved under a shorter day duration, then it is possible that the early-evolved algae had to synchronize their cell division cycle to shorter lengths of day than did later-evolved algae. We tested this hypothesis by growing Cyanobacteria, Dinophyceae, Prasinophyceae, Bacillariophyceae and Conjugatophyceae (evolutionary appearance probably in this order) at 8∶8 h light-dark cycles (LD), 10∶10 LD, and 12∶12 LD, at 20 or 27°C. Cyanobacteria synchronized their cell division cycles optimally at 8∶8 h LD, Dinophyceae and Prasinophyceae at 10∶10 h LD, and Conjugatophyceae and Bacillariophyceae at 12∶12 h LD. The synchrony of cell division was scarcely affected by temperature. Results suggested that the early evolved unicellular autotrophic organisms such as the Cyanobacteria synchronized their cell division cycle under a shorter day duration than later-evolved unicellular algae, and these traits may have been conserved by quiescent genes up to the present day.

A procedure to estimate okadaic acid in whole dinoflagellate cells using immunological techniques

A single procedure to detect and estimate okadaic acid in isolated whole cells was developed based on immunofluorescence and microscope photometry. This procedure allows the study of variations in okadaic acid concentration per cell although it is no substitute for HPLC procedures. Cells from mid-log exponential and stationary phase from two different clonal cultures of the okadaic-acid-producing dinoflagellate Prorocentrum lima (PI 5V and PI 7V) were analyzed. The results showed that: (1) cells from saturated phase cultures contain more okadaic acid than those from exponentially-growing mid-log phase; (2) genetic differences exist in okadaic acid production between the clones used; (3) okadaic acid is synthesized continuously during the whole cell cycle.

Early Development in Fucus spiralis: Analysis of Surface Glycan Moieties and Cell Differentiation Using Lectins

Glycan moieties of cell surfaces have been implicated as mediators in cell-cell interaction, cell differentiation and control of early development. At present, glycan moieties of cell surfaces are currently analyzed using lectins. Fucus spiralis is widely employed to study early development. In this work, the early development of Fucus spiralis embryos was analyzed using FITC-labelled lectins. The cell surfaces of spermatozoa showed more single patterns of glycan moieties than oocytes. However, just after fertilization young zygotes lose lectin binding sites suggesting important changes on their cell surface. Soon afterwards, zygotes polarize their surface glycans, and these show more diversity at rhizoid poles than at thallus poles (...)

Contact Inhibition: Also a Control for Cell Proliferation in Unicellular Algae?

According to traditional views, the proliferation of unicellular algae is controlled primarily by environmental conditions. But as in mammalian cells, other biological mechanisms, such as growth factors, cellular aging, and contact inhibition, might also control algal proliferation. Here we ask whether contact inhibition regulates growth in several species of unicellular algae as it does in mammalian cells. Laboratory cultures of the dinoflagellate Prorocentrum lima (Ehrenberg) Dodge show contact inhibition at low cell density, so this would be an autocontrol mechanism of cell proliferation that could also act in natural populations of P. lima. But, Synechocystis spp., Phaeodactylum tricornutum (Bohlin), Skeletonema costatum (Greville), and Tetraselmis spp. do not exhibit contact inhibition in laboratory cultures because they are able to grow at high cellular density. Apparently their growth is limited by nutrient depletion or catabolite accumulation instead of contact inhibition. Spirogyra insignis (Hassall) Kutz, Prorocentrum triestinum Schiller, and Alexandrium tamarense (Halim) Balech show a complex response, as they are able to grow in both low and high cell density medium. These results suggest that contact inhibition is more adaptative in benthic unicellular algae.

A Morphometrical Analysis of Ultrastructural Changes during the Cell Cycle of Prorocentrum triestinum Using Stereology

The ultrastructural changes of the dinoflagellate Prorocentrum triestinum Schiller were quantitatively analyzed during its cell cycle using stereological procedures on ultrathin serial sections and transmission electron microscopy. The absolute volume growth and volume variations of different structural parameters were calculated during a cell cycle. Cell organization was characterized using relative parameters: volume change of nuclei, chloroplasts, mitochondria, storage products and other organelles. Storage products were the cellular compartment with the highest degree of variation during the cell cycle. The changes in the total numbers of mitochondria and chloroplasts were also estimated

An apparent growth factor modulation of marine dinoflagellate excystment

Abstract The effects of growth factors on Alexandrium tamarense (Halim) Baleen excystment were analysed under laboratory conditions. The addition of 10 ng·ml−1 of platelet-derivated growth factor (PDGF) or 10% of fetal bovine serum (FBS), which are potent mitogens used to increase eukaryotic cell proliferation in culture, caused a statistically significant increase in excystment. While in the unsupplemented medium scarcely any excystment took place during the first days, most of the excystment in the supplemented medium took place during the first days of treatment with growth factors.

Characterization of morphospecies and strains of Pseudanabaena (Cyanophyceae) from laboratory cultures using antibodies and lectins

Immunofluorescence assays using polyclonal and preadsorbed antibodies as well as FITC-labelled lectins were employed to characterize two morphospecies of the cyanophytes Pseudanabaena catenata and P. planctonica from laboratory cultures isolated from two reservoirs in Spain and one in Denmark. Binding of both antibodies and lectins was unaffected by phase of the cell division cycle, growth phase and environmental factors such as culture medium, light or temperature. Good-quality polyclonal antibodies were obtained for several strains of both morphospecies. As expected, different morphospecies from the same location do not share common antigens while strains of the same morphospecies grown in different reservoirs do share most of their surface antigens. However, P. catenata grown at La Pinilla (Spain) does not share common antigens with P. catenata from Bastrup (Denmark). Geographical barriers, in this case, seem to play an important role in determining surface antigen diversity. Similar results are obtain...