Georgios Nikolaidis

Morphology, phylogeny and toxin analysis of Pseudo-nitzschia pseudodelicatissima (Bacillariophyceae) isolated from the Thermaikos Gulf, Greece

Moschandreou K.K., Papaefthimiou D., Katikou P., Kalopesa E., Panou A. and Nikolaidis G. 2010. Morphology, phylogeny and toxin analysis of Pseudo-nitschia pseudodelicatissima (Bacillariophyceae) isolated from the Thermaikos Gulf, Greece. Phycologia 49: 260–273. DOI: 10.2216/09-42.1 Pseudo-nitzschia pseudodelicatissima is a diatom species that has been associated with the production of the neurotoxin domoic acid (DA), causative agent of amnesic shellfish poisoning. However, the taxonomic revision of this species complex resulted in uncertain identity and, subsequently, toxicity of the strains tentatively identified under light microscope as P. pseudodelicatissima. Six strains isolated from the Thermaikos Gulf during the period from August to October 2008 were cultured and tested for production of DA. All strains were studied in morphological detail using electron microscopy and were identified as P. pseudodelicatissima. The identification was supported by phylogenetic analyses of the large subunit (LSU) and internal transcribed spacer (ITS) ribosomal rDNA regions, along with the secondary structure of ITS-2 rDNA region. DA was detected in all cultures and in three of the strains in quantifiable amounts up to 3.08 ng ml−1. Two of the strains were studied over the course of a batch culture growth experiment and DA was detected from the mid-exponential until late-stationary phases; the maximum whole-culture DA levels were measured in late-exponential to early-stationary phases. This study confirms the production of DA by P. pseudodelicatissima in culture.

Inter- and intra-specific diversity of Pseudo-nitzschia (Bacillariophyceae) in the northeastern Mediterranean

A total of 92 cultured Pseudo-nitzschia strains, established between March 2007 and April 2010 from Greek coastal waters, were identified by using morphological and molecular (ITS2 region) characters. Twelve species were identified, among which P. brasiliana, P. subpacifica and P. hasleana were detected for the first time near Greek and East Mediterranean coasts. One P. delicatissima-like morphotype and another strain that closely resembled P. dolorosa were also found. Morphology and ITS2 phylogeny indicated that the level of diversity within the genus is higher than previously estimated (e.g. among P. delicatissima-like diatoms). Additionally, the ITS2 phylogeny revealed genetic variation within species, indicative of geographical differentiation (in P. brasiliana, P. fraudulenta, P. arenysensis, P. delicatissima and P. galaxiae). The majority of the cultured strains were also tested for domoic acid (DA) production. Strains of only three species, P. pseudodelicatissima, P. galaxiae and P. pungens var. pungens, were found capable of producing DA.

Phytoplankton and physical-chemical features of Tavropos Reservoir, Greece

Phytoplankton biomass values in Tavropos Reservoir, ranging from 92 to 1071 mg m−3, are within a range characteristic of oligotrophic waters. The seasonal sequence of biomass shows three annual peaks, differing from the monoacmic pattern seen in oligotrophic lakes. This sequence was profoundly affected by changes in water withdrawal and inflow rates. Diatoms, cryptophytes, chrysophytes and dinoflagellates, in that order, were the major constituents of the reservoir phytoplankton. The succession, from diatoms and chrysophytes in late winter-spring, to centric diatoms in late spring-summer and again to diatom-chrysophytes in late autumn was similar to that in oligotrophic lakes.

Assessing the neurotoxic effects of palytoxin and ouabain, both Na⁺/K⁺-ATPase inhibitors, on the myelinated sciatic nerve fibres of the mouse: an ex vivo electrophysiological study.

Palytoxin (PlTX) is a marine toxin originally isolated from the zoantharians of the genus Palythoa. It is considered to be one of the most lethal marine toxins that block the Na⁺/K⁺-ATPase. This study was designed to investigate the acute effects of PlTX and ouabain, also an Na⁺/K⁺-ATPase blocker, on the mammalian peripheral nervous system using an ex vivo electrophysiological preparation: the isolated mouse sciatic nerve. Amplitude of the evoked nerve compound action potential (nCAP) was used to measure the proper functioning of the sciatic nerve fibres. The half-vitality time of the nerve fibres (the time required to inhibit the nCAP to 50% of its initial value: IT₅₀) incubated in normal saline was 24.5 ± 0.40 h (n = 5). Nerves incubated continuously in 50.0, 10.0, 1.0, 0.5, 0.250 and 0.125 nM of PlTX had an IT₅₀ of 0.06 ± 0.00, 0.51 ± 0.00, 2.1 ± 0.10, 8.9 ± 0.30, 15.1 ± 0.30 h, and 19.5 ± 0.20 h, respectively (n = 5, 3, 4, 4, 10). PlTX was extremely toxic to the sciatic nerve fibres, with a minimum effective concentration (mEC) of 0.125 nM (n = 5) and inhibitory concentration to 50% (IC₅₀) of 0.32 ± 0.08 nM (incubation time 24 h). Ouabain was far less toxic, with a mEC of 250.0 μM (n = 5) and IC₅₀ of 370.0 ± 18.00 μM (incubation 24.5 h). Finally, when the two compounds were combined--e.g. pre-incubation of the nerve fibre in 250.0 μM ouabain for 1 h and then exposure to 1.0 nM PlTX--ouabain offered minor a neuroprotection of 9.1-17.6% against PlTX-induced neurotoxicity. Higher concentrations of ouabain (500.0 μM) offered no protection. The mouse sciatic nerve preparation is a simple and low-cost bioassay that can be used to assess and quantify the neurotoxic effects of standard PlTX or PlTX-like compounds, since it appears to have the same sensitivity as the haemolysis of erythrocytes assay--the standard ex vivo test for PlTX toxicity.