Ángeles Moroño

Accumulation and transformation of DSP toxins in mussels Mytilus galloprovincialis during a toxic episode caused by Dinophysis acuminata.

The time course of several outbreaks of the diarrhetic shellfish poisoning (DSP) producer Dinophysis acuminata and the consequent kinetic of accumulation and loss of toxins in mussels Mytilus galloprovincialis feeding on them was studied. Samples of mussels and seawater were frequently (2-3 times a week) collected from a raft in the Ri;a de Vigo. DSP toxins content of mussels and water was analyzed by HPLC-FD and phytoplankton was quantified in an inverted light microscope. Only okadaic acid (OA) and some of its conjugated forms (OA CF), estimated by enzymatic hydrolysis, were found in the plankton samples obtained, comprised mainly of D. acuminata cells. The main accumulated form in mussels was OA reaching a maximum of 10.1 microg OA g(-1) in the digestive gland (d.g.) in 16 days, falling below the quarantine level (ca. 2 microg OA g(-1) d.g.) by 45 days. The low polarity conjugated forms (LPCF), estimated by hexane extraction, accounted for 6.2% of the total toxin burden of the mussels. To quantify the rates of the processes involved in the accumulation, transformation and loss of the toxins, two dynamic models, a one-compartment and a two-compartment, including OA and its conjugated forms as variables were designed and implemented. The one-compartment model provided a good fit to the OA and LPCF actual data (r(2)=0.92 and r(2)=0.94, respectively). The two-compartment model did not fit the data markedly better than its one-compartment counterpart (r(2)=0.93 and r(2)=0.95, for OA and LPCF, respectively). High hydrolysis rates were estimated for most of the OA CF, which means that these forms came largely from the ingested plankton. The low estimated acylation rates support the previous point and suggest that the formation of LPCF by direct acylation of the OA is of little importance in M. galloprovincialis. Only in cases where the intoxication period is very long, can the formed acyl-derivatives be important, because they seem to accumulate for a long time in the mussels, as suggested by the low hydrolysis and depuration rates estimated from model fitting.

The effect of mussel size, temperature, seston volume, food quality and volume-specific toxin concentration on the uptake rate of PSP toxins by mussels (Mytilus galloprovincialis Lmk)

The accumulation of paralytic shellfish poisoning (PSP) toxins by bivalves is a serious threat to public health all over the world. However, very little is known about the uptake kinetics of these toxins and the environmental factors that may modify this process. We have studied the effect of mussel size, temperature, seston volume, food quality, and volume-specific toxin concentration (VOSTOC), on the uptake rate of paralytic shellfish poisoning (PSP) toxins by mussels (Mytilus galloprovincialis), by means of a second order factorial experiment. Over a 3-day period, the mussels were fed artificial diets containing Alexandrium minutum AL1V (a PSP toxin producer), Tetraselmis suecica, Ensiculifera sp1 and silt, to the levels required by each treatment. Mussel size, seston volume and VOSTOC were found to be statistically significant when the total toxin accumulated per weight of wet tissue was considered. Mussel size affected the uptake negatively and latter two positively. The interactions, mussel size-VOSTOC and mussel size-food quality were also significant. The response was not linear as shown by the significance of the quadratic term of mussel size. Notwithstanding, when the PSP toxins accumulation per mussel was analysed, only one factor, the VOSTOC and the interactions, food quality-mussel size and food quality-seston volume, were found to be significant. VOSTOC was the most important factor in the accumulation of toxins, in our opinion, probably due to toxin assimilation being mainly regulated by the probability of contact between the toxins and the cellular walls of the digestive system. The size of the bivalve is also especially important because toxin concentration is usually calculated per weight of bivalve tissue and because the weight-specific ingestion increases with mussel size. The food quality, which was directly related to the assimilation of organic matter, had an inverse effect on toxin assimilation. In our opinion, this is probably due to the effect of inorganic particles in enhancing the disruption of Alexandrium cells. Temperature had no effect on the uptake rate except for the accumulation of the gonyautoxin GTX1.

Depuration of mussels (Mytilus galloprovincialis) contaminated with domoic acid

Abstract Domoic acid is a neurotoxin responsible for the Amnesic Shellfish Poisoning (ASP). With the aim of determining its depuration kinetics in raft mussels Mytilus galloprovincialis and the effects of body weight, salinity and temperature on it, an experiment involving these factors was carried out. Mussels which had incorporated this toxin to a concentration of 153 μg·g–1 of soft tissue during a bloom of the pennate diatom Pseudo nitzschia australis, were collected from the Galician Rias and placed at temperatures of 18 and 22 oC and salinities of 12.5 and 31. Mussel samples were taken at the start of the experiment and daily during the four subsequent days and the domoic acid contents of the soft tissues were analysed. The simple one-compartment kinetics, that is a kinetics in which all toxin depurates at the same rate, was fitted to the data, with good quantitative results. Notwithstanding, the deviations of the model from the actual data were dependent on time, suggesting that such a simple model is not enough to correctly describe the data, and that a more complex kinetics may be more adequate. A two-compartment kinetic model, in which two pools of toxins exist (compartments), each one depurating at different rate, described qualitatively better the depuration but its quantitative contribution to the fit was not statistically significant. The parameters of the model, obtained by least squares fitting, suggest the possibility of a small second compartment of very small or null depuration rate, as detected in other species. This kind of model would explain the reduced quantitative contribution of the second (slowly depurating) compartment.Low salinity was shown to reduce the depuration rate. The two other factors checked, temperature and body weight, nor any interaction had significant effect on depuration rate.

Evaluation of passive samplers as a monitoring tool for early warning of dinophysis toxins in shellfish

From June 2006 to January 2007 passive samplers (solid phase adsorbing toxin tracking, SPATT) were tested as a monitoring tool with weekly monitoring of phytoplankton and toxin content (liquid chromatography–mass spectrometry, LC-MS) in picked cells of Dinophysis and plankton concentrates. Successive blooms of Dinophysis acuminata, D. acuta and D. caudata in 2006 caused a long mussel harvesting closure (4.5 months) in the Galician Rías (NW Spain) and a record (up to 9246 ng·g resin-week−1) accumulation of toxins in SPATT discs. Best fit of a toxin accumulation model was between toxin accumulation in SPATT and the product of cell densities by a constant value, for each species of Dinophysis, of toxin content (average) in picked cells. Detection of Dinophysis populations provided earlier warning of oncoming diarrhetic shellfish poisoning (DSP) outbreaks than the SPATT, which at times overestimated the expected toxin levels in shellfish because: (i) SPATT accumulated toxins did not include biotransformation and depuration loss terms and (ii) accumulation of toxins not available to mussels continued for weeks after Dinophysis cells were undetectable and mussels were toxin-free. SPATT may be a valuable environmental monitoring and research tool for toxin dynamics, in particular in areas with no aquaculture, but does not provide a practical gain for early warning of DSP outbreaks.

Effect of the industrial steaming on the toxicity, estimated by LC–MS/MS, of mussels exposed for a long time to diarrhetic shellfish poisoning (DSP) toxins

The effect of industrial steaming on mussels that had been naturally exposed to DSP toxins for a long time was studied using LC-MS/MS. The estimated toxicity increased with steaming by a percentage that cannot be explained by weight loss. The estimated toxin content per mussel increased substantially with the treatment, which can only be explained by an incorrect estimation by the technique (at the extraction or analytical level) or by the presence of unknown derivatives or analogues. Direct alkaline hydrolysis of the mussel meat yielded more toxin than the standard hydrolysis (hydrolysis of the methanolic extracts), suggesting that extraction was, at least in part, responsible for the increase of toxin content. In situations as the one described in this work, it can be expected that mussels with toxicities well below the regulatory limit could easily surpass that level after industrial steaming, thus producing important losses for food processors.

Presence of azaspiracids in bivalve molluscs from Northern Spain

ABSTRACT Low concentrations of azaspiracids have been found in bivalve molluscs all over the Northern and Northwestern coast of the Iberian Peninsula. The detections started in June 2016 and lasted until March 2017. The observed toxin profile was dominated by AZA2, followed by AZA1 and some other AZAs that were detected only in some samples. Some compounds producing fragments characteristics of AZAs but that do not fit with any of the known ones were also found. The causative agent has not been identified but, in sight of the toxin profile in the bivalves, it seems that it should be a new species or strain. The detections of AZAs in bivalves in the Northern Coast was linked to downwelling or upwelling relaxation and, in the Galician Rías, took place (with only a few exceptions) in the outer (more oceanic) part, suggesting that the responsible species develops at the open sea and that the populations are advected to the shore. HIGHLIGHTSAzaspiracids are present in bivalves from the north and northwest coast of Spain.AZA2 was the dominant AZA in the bivalves.Other AZAs as AZA 1, 3, 6 and 43 were found in some samples.The producer species seems to develop offshore.

Effect of the industrial canning on the toxicity of mussels contaminated with diarrhetic shellfish poisoning (DSP) toxins

The effect of canning in pickled sauce and autoclaving on weight, toxin content, toxin concentration and toxicity of steamed mussels was studied. Weight decreased by 25.5%. Okadaic acid (OA) and DTX2 content of mussel meat decreased by 24.1 and 42.5%, respectively. The estimated toxicity of the mussel remained nearly unchanged (increased by 2.9%). A part of the toxins lost by the mussels was leached to the sauce but the remaining part should have been thermally degraded. DTX2 underwent more degradation than OA and, in both toxins, free forms more than conjugated ones. This process, therefore, cannot be responsible for the large increments of toxicity of processed mussels -relative to the raw ones-sometimes detected by food processing companies. The final product could be monitored in several ways, but analysing the whole can content or the mussel meat once rehydrated seems to be the most equivalents to the raw mussel controls.