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Ubiquitous 'benign' alga emerges as the cause of shellfish contamination responsible for the human toxic syndrome, azaspiracid poisoning.

A new human toxic syndrome, azaspiracid poisoning (AZP), was identified following illness from the consumption of contaminated mussels (Mytilus edulis). To discover the aetiology of AZP, sensitive analytical protocols involving liquid chromatography-mass spectrometry (LC-MS) were used to screen marine phytoplankton for azaspiracids. Collections of single species were prepared by manually separating phytoplankton for LC-MS analysis. A dinoflagellate species of the genus, Protoperidinium, has been identified as the progenitor of azaspiracids. Azaspiracid-1, and its analogues, AZA2 and AZA3, were identified in extracts of 200 cells using electrospray multiple tandem MS. This discovery has significant implications for both human health and the aquaculture industry since this phytoplankton genus was previously considered to be toxicologically benign. The average toxin content was 1.8 fmol of total AZA toxins per cell with AZA1 as the predominant toxin, accounting for 82% of the total.

Animals↗

Occurrence of paralytic shellfish poisons in Thai freshwater puffers.

Screening tests were carried out on the toxicity of freshwater puffers Tetraodon leiurus complex and Tetraodon suvatii collected from Udonthani province, north-eastern Thailand. Toxicity was highest in the liver and varied according to the location and season of fish catch. Fish which were reared in tap water for 3 months reduced the toxicity substantially. Partial purification was achieved by an ultrafiltration technique. Toxin components were consequently identified by high-performance liquid chromatography. It was found that toxins separated from the eggs, liver, skin and muscle of these puffers were composed of saxitoxin, neosaxitoxin and decarbamoylsaxitoxin.

Animals↗

Invertebrate cell culture considerations: insects, ticks, shellfish, and worms.

Establishment of cell lines from insect and arachnid invertebrates has become routine, whereas other invertebrate taxa have been frustratingly unproductive of cell lines. None is available for any marine invertebrate, despite a strong and well-recognized need for cell lines from species that are important in aquaculture, from parasite vectors and intermediate hosts of parasites, from parasites themselves, from certain biomedical models, and from other species that are pests. Drawing on experiences gained attempting to establish cell lines from molluscs and trematodes and on published and ongoing research with diverse invertebrates, this chapter attempts to anticipate the problems that are likely to be encountered in such endeavors and discusses possible solutions. Criteria to be considered in the selection of basic culture media, temperature, pH, and media additives; approaches that have been developed to yield sterile primary cultures; and factors to consider in decisions about feeding schedules, retention of tissue fragments and nonadherent cells, use of heterologous feeder layers, and other variables are described. Suggestions are made concerning means to objectively score the success of tested variables and means to induce cell replication. The chapter ends with notes on conventional means to characterize cell lines and an account of contemporary efforts to immortalize cells by means of genome manipulation. Enduring success with a single molluscan cell line, transient successes with crustacean and helminth cell lines, and promising developments in transgenesis with invertebrates all lead to the hopeful conclusion that the invisible barrier to cell propagation in historically refractory species will soon be a thing of the past.

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Transfer constants for blood-brain barrier permeation of the neuroexcitatory shellfish toxin, domoic acid.

The cause of the toxic mussel poisoning episode in 1987 was traced to a plankton-produced excitotoxin, domoic acid. Experiments were undertaken to quantitate the degree to which blood-borne domoic acid can permeate the microvasculature to enter the brain. Pentobarbital-anesthetized, adult rats received an i.v. injection of 3H-domoic acid which was permitted to circulate for 3-60 min. Transfer constants (Ki) describing blood-to-brain diffusion of tracer were calculated from analysis of the relationship between brain vs plasma radioactivity with time. Mean values (mL.g-1.s-1 X 10(6] for permeation into 7 brain regions (n = 10 rats) ranged from 1.60 +/- 0.13 (SE) to 1.86 +/- 0.33 (cortex, pons-medulla respectively), and carrier transport or regional selectivity in uptake were not evident. Nephrectomy prior to domoic acid injection resulted in the elevation of circulating plasma tracer level and brain uptake. The Ki values are comparable to those for other polar compounds such as sucrose, and indicate that the blood-brain barrier greatly limits the amount of toxin that enters the brain. Together with absorbed dosage, integrity of the cerebrovascular barrier and normal kidney function are important to the outcome of accidentally ingesting domoic acid.

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Solubility improvement of shellfish muscle proteins by reaction with glucose and its soluble state in low-ionic-strength medium.

When myofibrillar proteins of scallop striated adductor muscle were reacted with glucose through the Maillard reaction, the change in the solubility of myofibrillar proteins in 0.05-0.5 M NaCl solutions during glycosylation and their soluble states were investigated. The solubility in low-ionic-strength media increased greatly with the progress of the Maillard reaction. The solubility in 0.1 M NaCl reached 83% when more than 60% of lysine residues in myofibrillar proteins were modified by glucose. However, the excess progress of the Maillard reaction impaired the improved solubility of myofibrillar proteins in a low-ionic-strength medium. Myosin, actin, and paramyosin in glycosylated myofibrillar proteins were solubilized independently regardless of NaCl concentration. In addition, the glycosylated myosin lost its filament-forming ability and existed as a monomer in 0.1 M NaCl.

Actins↗

Analyses of glycolipids from fish, shellfish, and sea snake lipids by high-performance liquid chromatography.

To determine the existence of glycolipids (neutral glycosphingolipid and glycoglycerolipid) in sea snake, round frigate mackerel, sardine, sea urchin, and abalone, we performed silica gel chromatography and high-performance liquid chromatography (HPLC) using an Aquasil-SS column and a C(8)-reversed phase silica gel column. HPLC with a UV absorption detector was used to analyze neutral glycosphingolipid. These chromatograms showed typical peaks in round frigate mackerel lipid, in sea snake crude fat, in abalone intestine lipid, and in sea urchin intestine lipid. UV-HPLC was also used to analyze glycoglycerolipid. These chromatograms indicated a large peak in round frigate mackerel lipid and a small peak in purified sardine oil. In addition, we observed the same peaks in the glycolipid fraction of round frigate mackerel muscle lipids and sea snake crude fat using a differential refractometer detector. The results of this study suggest that the peaks are neutral glycosphingolipid or glycoglycerolipid and that neutral glycosphingolipid and glycoglycerolipid may have specific physiological functions in each living creature.

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