Do toxic Pseudo-nitzschia species pose a threat to aquaculture in the southern Benguela eastern boundary upwelling system?

The productive but highly exposed coastline of the southern Benguela eastern boundary upwelling system offers limited natural environment for aquaculture. Saldanha Bay located on the west coast of South Africa is one of the few embayments on the coastline that provides a productive and relatively sheltered environment suitable for the cultivation of shellfish. Consequently, bivalve culture in South Africa is centered in Saldanha Bay and is presently targeted for expansion. Pseudo-nitzschia blooms including toxin-producing species are shown to contribute significantly to the phytoplankton of Saldanha Bay specifically in spring and summer. Their dominance at this time of the year, when upwelling is strongest, fits the ecological profile of Pseudo-nitzschia occurring during periods of high turbulence and nutrients. Multiple Pseudo-nitzschia blooms were sampled under varying environmental conditions and the strength of the relationship between Pseudo-nitzschia cell abundance and particulate domoic acid (pDA) content, reflecting bloom toxicity, varied greatly. This variability is the result of the combined influence of species and strain composition of the Pseudo-nitzschia assemblage and the effect of environmental conditions on toxin production. Elevated levels of pDA were associated with higher concentrations of cells of the P. seriata complex differentiated by frustule width (>3 µm). P. australis was identified as a toxin-producing species and a prominent member of the P. seriata complex. Low DA levels in shellfish in Saldanha Bay are considered a function of low cellular domoic acid (cDA). Silicate limitation has emerged as an important factor inducing DA production in Pseudo-nitzschia species. The high ratio of silicate to nitrate in Saldanha Bay provides a plausible explanation for the low toxin content of Pseudo-nitzschia blooms in the bay and the consequent low risk posed by these blooms to the aquaculture sector.

Latitudinal transects in the southeastern Pacific Ocean reveal a diverse but patchy distribution of phycotoxins.

Phycotoxin distribution and abundance was determined during an oceanographic expedition along a latitudinal transect of 27° extent in the southeastern Pacific Ocean, from the fjords of Tierra del Fuego Island to offshore Copiapó in the Atacama region along the Chilean coast. Plankton samples were harvested at regular intervals during the entire cruise and later analyzed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for domoic acid (DA) and lipophilic toxins. Although no evident toxic algal bloom was encountered during this transect, several phycotoxin analogues from distinct toxin groups were detected. These phycotoxins included DA, the pectenotoxins PTX-2, PTX-2sa and PTX-11, dinophystoxin-1 (DTX-1) and gymnodimine (GYM), which is the first report of this latter toxin in the southeast Pacific. A region-specific and rather disjunct distribution of GYM, DA and DTX-1 was observed, whereas PTX-2, PTX-2sa and PTX-11 were more widely distributed over almost the entire transect. This work represents the first assessment of lipophilic toxins through a wide latitudinal transect of the southeastern Pacific, revealing a patchy distribution of several phycotoxins and pointing out the specific geographical distribution of the putative toxigenic organisms.

Characterization of spirolides a, c, and 13-desmethyl c, new marine toxins isolated from toxic plankton and contaminated shellfish.

Three additional marine toxins, spirolides A (1), C (3), and 13-desmethyl-C (7), were isolated from contaminated scallops and phytoplankton collections obtained from a Nova Scotian aquaculture site, as well as from batch cultures of the dinoflagellate Alexandrium ostenfeldii obtained as a single-cell isolate from these phytoplankton assemblages. The structures of these new spirolide derivatives, characterized by mass spectrometry and NMR, indicate a close relationship with spirolides B (2) and D (4) isolated previously from contaminated shellfish in the same area. All of these compounds display "fast-acting" toxicity in the traditional bioassay used for monitoring shellfish, and this is related to the presence of a cyclic imine function in all these compounds. Those spirolides containing a vicinal dimethyl group in the seven-membered ring are resistant to oxalic acid hydrolysis, whereas those that do not are readily hydrolyzed. These observations suggest that the extra methyl group on the seven-membered imine ring of 3, 4, and 7 appears to block the process of imine hydrolysis perhaps by stereochemical interference.

Spirolide composition of micro-extracted pooled cells isolated from natural plankton assemblages and from cultures of the dinoflagellate Alexandrium ostenfeldii.

A novel micro-extraction technique was applied to the extraction of biologically active macrocyclic imines known as spirolides from pooled individual cells isolated from spirolide-rich plankton material. For comparison, this method was also applied to pooled individual cells isolated from a unialgal culture of the marine dinoflagellate Alexandrium ostenfeldii (Paulsen) Balech & Tangen, a species known to produce spirolides. Both athecate cells and motile forms of gonyaulacoid dinoflagellates derived from size-fractionated plankton material from Nova Scotia, Canada were sorted and pooled by the glass micropipette isolation technique and by flow cytometry. The development of a highly sensitive analytical method for spirolides (detection limit 2 ng ml(-1) for spirolide B) using liquid chromatography-mass spectrometry (LC-MS) and application to micro-extracted samples allowed the accurate determination of spirolide composition in as few as 50 cells. Total spirolide concentrations (fmol cell(-1)) calculated from pooled micropipette isolated cells were very consistent with those based upon bulk- or micro-extractions of A. ostenfeldii cells from unialgal batch cultures in exponential growth phase. The results of the pooled cell selection from field material from two sites in Nova Scotia confirmed the association of spirolides with vegetative cells of A. ostenfeldii and related athecate forms. Combining these techniques represents a highly sensitive method for the analysis of marine toxins within complex plankton matrices, even when the toxigenic species is in low abundance, by enrichment of the target organism.

Cross-reactivity of an anti-okadaic acid antibody to dinophysistoxin-4 (DTX-4), dinophysistoxin-5 (DTX-5), and an okadaic acid diol ester.

The cross-reactivity of the 6/50 monoclonal anti-okadaic acid antibody (mAb) to the recently discovered diarrhetic shellfish poisoning (DSP) metabolites dinophysistoxin-4 (DTX-4), dinophysistoxin-5 (DTX-5), and an okadaic acid (OA) diol ester was determined using a competitive indirect enzyme-linked immunosorbent assay (ELISA). The reactivity of the antibody to these molecules was compared to that with OA; the 6/50 mAb recognized all of these DSP compounds with equal sensitivity within the working range of the antibody (10-100 nM for OA). This confirms the ability of the antibody to detect all DSP compounds when used in analyses including ELISA and immunocytochemistry.

Environmental stress and domoic acid production by Pseudo-nitzschia: a physiological perspective.

Production of domoic acid (DA) by the pennate diatom Pseudo-nitzschia multiseries is associated with physiological stress caused by silicate (Si) and/or phosphate (P) limitation. Such limitation may promote DA synthesis by (1) reducing primary metabolic activity, thus making available necessary precursors, high energy compounds, and cofactors, and (2) favoring the expression of genes involved in the biosynthesis of this toxin. In the case of Si and P-limitation, DNA synthesis and the progression through the cell division cycle are slowed, perhaps prolonging or arresting the cells in the stage of the division cycle which is most conducive to DA production. However, N-limitation results in an insufficient pool of cellular free N, which restricts synthesis of this nitrogenous toxin. A continuous supply of photophosphorylated high-energy intermediates (e.g., ATP and NADPH) is necessary for DA synthesis. In order to better understand the mechanism(s) of DA production, more studies are needed to elucidate: (1) the details of the biosynthetic pathway, (2) the regulation of enzymes involved in the pathway, (3) the relation between DA synthesis and the cell division cycle, (4) the cellular compartmentalization of DA biosynthesis, and (5) other environmental factors that may trigger DA production. Finally, these studies should be extended to include toxigenic Pseudo-nitzschia species other than P. multiseries, to confirm the commonality of these mechanisms.

Accumulation of paralytic shellfish toxins by surfclams, Spisula solidissima (Dillwyn, 1897) in the Gulf of Maine: seasonal changes, distribution between tissues, and notes on feeding habits.

Accumulation of paralytic shellfish poisoning (PSP) toxins by surfclams, Spisula solidissima, was studied over a period of two years at two inshore locations in southern Maine and at six stations on Georges Bank in the Gulf of Maine. Whole animals as well as individual tissues (siphon, mantle, digestive gland, foot, adductor muscle, gill) were analyzed for PSP-toxicity levels using the standard AOAC mouse bioassay. Analyses of gut contents were carried out on surfclams from both inshore and offshore locations to identify the type of particles ingested. Surfclams feed primarily on phytoplankton and detrital material characteristic of the overlying seawater and surface sediment. No evidence was found for any selection based on particle size or type. Elevated levels of PSP toxins were noted in surfclams from Georges Bank more than two years after initial toxification. Toxins were not evenly distributed among the various tissues of surfclams. Initially, maximum toxicity among surfclam tissues was found in digestive glands; however, subsequent analyses of samples collected later in the year indicated that toxicity in gill and mantle tissues had increased relative to initial values. No toxicity was detected in adductor muscles. Surfclams are characterized by a high variation in total toxin load among individual animals, with a tendency for decreasing variation as toxin levels increase. Archived data from the Main Department of Marine Resources revealed annual and seasonal patterns of toxin accumulation by surfclams, i.e., toxin accumulation is an annual event, with initial increases in toxicity usually occurring in early spring.

The utilization of inorganic and organic phosphorous compounds as nutrients by eukaryotic microalgae: a multidisciplinary perspective: part 1.

This comprehensive literature review of the phosphorus nutrition and metabolism of eukaryotic microalgae deals sequentially with (1) extracellular P-compounds available for algal utilization and growth; (2) orthophosphate uptake mechanisms, kinetics, and influence from environmental variables; (3) phosphatase-mediated utilization of organic phosphates involving multiple enzymes, induction and cellular location of repressible and irrepressible phosphatases, and their role in growth physiological processes; (4) intracellular phosphate metabolism covering diversity of phosphometabolites. ATP-linked energy regulation, polyphosphate pools and storage roles, phospholipids and phospholipases; (5) steady-state and transient-state models relating phosphate utilization to growth; (6) ecological aspects covering manifestations of phosphorus limitation, interspecific competition for phosphonutrients among microorganisms, and current views on phosphorus cycling and turnover in aquatic ecosystems. Although concentrating on the microalgae, the review often points out sounder conclusions drawn from bacteria and fungi, and includes specific macroalgae in considering certain subtopics where such algae were better investigated and provided a good basis for comparison with the microalgae.