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Distribution of paralytic toxins in California shellfish.

Samples of Saxidomus nuttali and Mytilus californianus collected during the 1981 dinoflagellate bloom at Bodega Bay, California, were analyzed for the presence of paralytic toxins. Neck tissue of S. nuttali contained saxitoxin (STX) and neoSTX (95% of the total toxicity), whereas the bodies contained neoSTX and a mixture of the gonyautoxins. In a sample of M. californianus the presence of neoSTX and the gonyautoxins was demonstrated, whereas a second sample, collected at a different site, contained almost exclusively neoSTX.

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Production of paralytic shellfish toxins by a bacterium Moraxella sp. isolated from Protogonyaulax tamarensis.

A bacterium Moraxella sp. isolated from Protogonyaulax tamarensis was cultured in various conditions. Changes of toxicity and toxin components of the cells during culture were analyzed by bioassay and HPLC-fluorometric analysis. Toxin productivity of Moraxella sp. increased when it was cultured in nutrition-deficient environments. The main toxins produced by Moraxella sp. in these conditions were gonyautoxins (GTXs), mainly GTX 1 and 4 which are major toxins of P. tamarensis.

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Diarrhoeic shellfish toxins in Adriatic Sea mussels evaluated by an ELISA method.

A competitive enzyme immunoassay (ELISA) was used to determine the presence of okadaic acid (OA) and/or dinophysistoxin-1 (DTX-1) in mussels. The accuracy and sensitivity of the ELISA method has been checked. The sensitivity of the method (100 ng/g of hepatopancreas) makes it possible to determine OA concentrations ten times lower than the tolerance limits established by the Health Authorities of many countries. For the first time, OA and/or DTX-1 were detected in mussels (Mytilus galloprovincialis) collected in different stations along the Adriatic Sea coasts.

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Comparison of carotenoids in the ovaries of marine fish and shellfish.

1. The following carotenoids were isolated and identified: astaxanthin diester, tunaxanthin monoester, astaxanthin monoester, tunaxanthin, astaxanthin, doradexanthin, lutein, zeaxanthin, idoxanthin, triol and tetrol from nine species of fish; astaxanthin diester, astaxanthin monester, astaxanthin, doradexanthin, zeaxanthin, idoxanthin and tetrol from four species of crustacean, astaxanthin, pectenolone, pectenoxanthin, pectenol and tetrol from four species of scallop. 2. Tunaxanthin monoester and astaxanthin diester were major carotenoids in skipjack and Pacific cod, respectively. 3. The concentration of carotenoids ranged 0.065-1.95, 1.30-5.91 and 1.56-7.15 mg per 100 g ovary for fish, crustacean and scallop, respectively. 4. The species- and tissue-specificity of ovarian carotenoids and their possible role are discussed.

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Accumulation, transformation and tissue distribution of domoic acid, the amnesic shellfish poisoning toxin, in the common cuttlefish, Sepia officinalis.

Domoic acid (DA) is a phycotoxin produced by some diatoms, mainly from the Pseudo-nitzschia genus, and has been detected throughout the marine food web. Although DA has been frequently found in cephalopod prey such as crustaceans and fish, little is known about DA accumulation in these molluscs. This study presents the first data showing relevant concentrations of DA detected in the common cuttlefish, Sepia officinalis, which is one of the most studied cephalopod species in the world. Domoic acid was consistently found throughout 2003 and 2004 in the digestive gland of cuttlefish reaching concentrations of 241.7 microg DA g(-1). The highest DA values were detected during spring and summer months, periods when Pseudo-nitzschia occur in the plankton. In fact, Pseudo-nitzschia blooms preceded the highest DA concentrations in cuttlefish. Evaluation of DA tissue distribution showed elevated DA concentrations in the digestive gland and branchial hearts. Further, DA isomers comprised a relevant percentage of the toxin profile, indicating degradation and biotransformation of the toxin in the branchial hearts. The common cuttlefish, like other cephalopod species, plays a central position in the food web and might be a new DA vector to top predators like marine mammals. Human intoxications are not expected since DA was only seldom detected in the mantle and even then in very low levels (max 0.7 microg DA g(-1)). However, in some countries whole juvenile animals are consumed (i.e. without evisceration) and in this case they might represent a risk to human health. This study contributes to understanding the occurrence of phycotoxins in cephalopods and reveals a new member of the marine food web able to accumulate DA.

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Bioconversion of shellfish chitin wastes for the production of Bacillus subtilis W-118 chitinase.

Bacillus subtilis W-118, a strain that produces antifungal materials, excreted a chitinase when cultured in a medium containing shrimp- and crab-shell powder as the major carbon source. This chitinase, purified by sequential chromatography, had a molecular mass of 20,600 Da and a pI of 6. The optimum pH, optimum temperature, and pH stability of the chitinase were pH 6, 37 degrees C, and pH 5-7, respectively. The unique characteristics of the purified chitinase include low molecular mass and acidic pI. In the investigation of the inhibitory activity, it was found that the growth of Fusarium oxysporum was 100% inhibited after incubation for 1 day with sterilized W-118 chitinase solution (5.6 units/mL). The chitinase hydrolyzates of chitin with low degrees of polymerization (DP 1-6) were analyzed by HPLC. Longer reaction times led to the generation of chitin oligosaccharides with lower DP. The chitin oligosaccharides were examined for their inhibitory effects on F. oxysporum and human leukemia cell lines.

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Determination of putrescine and cadaverine in seafood (finfish and shellfish) by liquid chromatography using pyrene excimer fluorescence.

A liquid chromatography (LC) method is described for the easy determination of the biogenic diamines putrescine (PUT) and cadaverine (CAD) in canned tuna, frozen tuna loin, fresh mahimahi fillet, frozen raw shrimp, cooked lump crabmeat, and fresh and cold-smoked salmon. The method is also a useful screen for histamine (HTA). The method involves homogenization of fish tissue, extraction of biogenic amines into borate-trichloroacetic acid solution, centrifugation, and derivatization of supernatant with 1-pyrenebutanoic acid succinimidyl ester. The derivatized diamine species allow for the intramolecular excimer fluorescence of the pyrene moiety at a higher emission wavelength than is possible for the endogenous tissue monoamines, thus providing visual specificity of detection. All seafood species were fortified with 0.5, 1.0, 5.0, 10.0, and 15.0 microg/g (ppm) of PUT and CAD. Determination was based on standard graphs for PUT and CAD using peak areas with standard solutions equivalent to 0.375, 1.0, 5.0, 10.0, and 20.0 ppm in tissue. A set of five matrix controls (unfortified seafood tissue) were also analyzed; endogenous PUT was found in all samples except the canned tuna, and CAD found only in the shrimp, crab, and cold-smoked salmon. The background amines were thus subtracted prior to determining spike recovery. The intra-assay average recoveries ranged from 71 to 94% across species and spike levels.

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Accumulation of organic and inorganic contaminants in shellfish collected in estuarine waters near Pensacola, Florida: contamination profiles and risks to human consumers.

We conducted a screening level assessment of contaminants in blue crabs (Callinectes sapidus) and oysters (Crassostrea virginica) from bays and bayous in the Pensacola, FL area. Tissue samples were analyzed for 17 dioxins/furans, 12 dioxin-like PCB (DL-PCBs) congeners, mercury, and various metals. Contaminant levels were compared to screening values (SV) calculated using U.S. EPA recommendations for establishing consumption advisories. All sampling locations exceeded the SV (0.098pgg(-1)) for dioxins/furans/DL-PCBs, based on a Florida-specific consumption rate (46gday(-1)). Arsenic (inorganic), mercury, cadmium, and zinc levels exceeded SVs in samples from select locations, and with the exception of mercury, these locations were generally downstream of known contaminated areas. We also assessed potential human health risks from consumption of these species. Risks to human health were greatest from consumption of crab hepatopancreas, suggesting that consumption of hepatopancreas, whether directly or indirectly, from crabs collected anywhere in the Pensacola Bay region should be avoided.

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Okadaic acid, a causative toxin of diarrhetic shellfish poisoning, in green-lipped mussels Perna viridis from Hong Kong fish culture zones: method development and monitoring.

Green-lipped mussels (Perna viridis) were collected from seven fish culture zones (FCZs) in Hong Kong and analyzed for okadaic acid (OA). A conventional HPLC method was modified by incorporating a proteinase K digestion step. Results suggest that a higher recovery (2.5 times higher) of OA was obtained from spiked samples after the addition of 1.08 mg proteinase K in comparison with samples incubated without the proteolytic enzyme. For the hepatopancreas (HP) of individual field-collected mussels, the additional digestion step can enhance OA extraction by 3.1 times. Spatial and temporal variations in OA concentrations in the mussels from various FCZs were investigated. The highest concentration of OA in mussel HP samples was 1164.9 ng/g HP wet wt. With respect to OA concentrations in whole mussel tissues from seven sites and four seasonal samplings, the concentrations were between 70.0 and 131.0 ng/g wet wt., which did not exceed the generally recognized international regulatory criteria (>200 ng/g) for OA.

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Remarkable difference in accumulation of paralytic shellfish poisoning toxins among bivalve species exposed to Pyrodinium bahamense var. compressum bloom in Masinloc Bay, Philippines.

Seasonal variation of bivalve toxicity was monitored in association with the abundance of the toxic dinoflagellate Pyrodinium bahamense var. compressum in Masinloc Bay, Luzon Island. Among 7 species of bivalve, 6 species became toxic during a bloom of the dinoflagellate. However, remarkable difference in the toxicity was observed among the species. The toxicity of thorny oyster Spondylus squamosus was the highest among the species, showing more than 30 times that of safety consumption level after the peak bloom of the dinoflagellate, while other bivalve species showed much lower toxicity. The toxicity of thorny oyster decreased under absence of the dinoflagellate, but this species maintained a considerably high toxicity throughout a year. Similar trend was observed in penshell Atrina vexillum in a small scale, indicating that these species in the bay are not safe for human consumption almost throughout a year. The toxicity of green mussel Perna viridis increased to significant level during a bloom, but it decreased rapidly when the dinoflagellate disappeared. Toxin analysis of cultured and natural cells showed typical toxin profile of the dinoflagellate. Toxin profile of all the bivalve species reflected the characteristic toxin profile of the dinoflagellate.

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Giardia in shellfish-farming areas: detection in mussels, river water and waste waters.

Giardia cyst contamination of mussels (Mytilus galloprovincialis), raw and treated waste waters and water from rivers that flow into four Galician estuaries (NW Spain), where bivalve molluscs are cultured for human consumption, was studied. The high prevalence of contamination in mussels (41.8%, n=184), raw waste water (90.9%, n=11), treated waste water (87.5%, n=16) and in samples of river water (85.7%, n=7), with cyst counts of 9.8-1800.0, 7.0-2541.0 and 1.0-29.3 cysts l(-1), respectively, illustrate the wide distribution of this enteropathogen in the environment and the potential risk to public health associated with the consumption of raw or undercooking bivalves and use of these estuaries for recreational purposes.

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