Spatial and temporal evolution of PSP toxins along the Atlantic shore of Morocco.

A monitoring program for bivalve molluscs contaminated by algal toxins was established in 1992 at different stations along the Atlantic Moroccan shore. The presence of toxicity in bivalve molluscs commercially exploited was tested fortnightly using the mouse bioassay method. Results obtained from this surveillance indicate paralytic shellfish poisoning is responsible for bivalve molluscs contamination along the Atlantic coastline of Morocco. Toxin profile was established by automated pre-column HPLC/FLD in selected contaminated tissues. The study of individual toxins in mussel during a bloom in the northern Atlantic coastline in 1994 showed a very complex profile, typical to that obtained with cultures of the toxigenic dinoflagellate Gymnodinium catenatum isolated from the Iberian region. However, toxin composition of mussels and marine beans from later blooms in the southern Atlantic coastline in 1999 showed a strong resemblance with that of Alexandrium minutum, due to dominance of gonyautoxins 1/4. A minor contamination by G. catenatum due to the presence of decarbamoyl-saxitoxin is hypothesized.     

Detection of paralytic shellfish poisoning (PSP) toxins in shellfish tissue using MIST Alert, a new rapid test, in parallel with the regulatory AOAC mouse bioassay.

In parallel trials with the mouse bioassay, MIST Alert for Paralytic Shellfish Poisoning (PSP), a rapid diagnostic test for PSP, detected 100% of the toxic extracts in over 2100 regulatory samples. Toxic extracts contained at least 80 microg saxitoxin equivalents (STX equiv.) in 100 g of shellfish tissue, or more, as measured by the regulatory AOAC mouse bioassay. Only one potentially toxic sample, which contained 78 and 86 microg STX equiv./100 g shellfish tissue in two different mouse bioassays, was recorded as negative in one replicate of MIST Alert. All other toxic extracts among more than 2100 regulatory shellfish tissue samples were detected by MIST Alert for PSP. The MIST Alert for PSP also detected the majority of extracts containing PSP toxin greater than 32 microg STX equiv./100 g, which is the mouse bioassay detection limit. The MIST Alert for PSP gave a false positive result compared to the mouse bioassay at an average rate of about 14% over all sites, although some differences were seen between sites. Further analysis by high performance liquid chromatography (HPLC) of the (false positive) extracts showed that many contained PSP toxicity in the range of 20-40 microg STX equiv./100 g, below the level detectable by the mouse bioassay. The MIST Alert for PSP gave false positive results from extracts containing less than 20 microg STX equiv./100 g shellfish tissue only about 6% of the time. The PSP family of toxin analogues can occur in any combination in naturally contaminated shellfish tissue and the antibody mixture in the MIST Alert tests detect each of the different PSP toxin analogues with different efficacy. It is therefore impossible to provide an exact detection limit for the MIST Alert that would be applicable for all possible toxin profiles. Through the experience of comparison testing with the regulatory mouse bioassay in many parts of the world, with over 2100 different samples, the MIST Alert for PSP has proven its ability to detect all types of profiles of the PSP toxin analogues. The detection limit for MIST Alert for PSP was about 40 microg STX equiv./100 g for the 'average' profile of PSP toxin analogues. Since the detection limit depends on the toxin profile in the individual extract, it will also vary depending on the profile of analogues most commonly found at each geographic location. This was observed in our study. Over all sites in the trials, approximately 5% of samples below 40 microg STX equiv./100 g were positive, and 5% of samples between 40-80 microg STX equiv./100 g were negative. This is a reflection of the different analogue profiles found in naturally contaminated extracts, even after acid hydrolysis using the AOAC extraction method.     

Comparative study on differential accumulation of PSP toxins between cockle (Acanthocardia tuberculatum) and sweet clam (Callista chione).

At the western Mediterranean coast of Morocco, the cockle (Acanthocardia tuberculatum) contained persistent high levels of paralytic shellfish toxins for several years, while other bivalve molluscs such as sweet clam (Callista chione) from the same vicinity were contaminated seasonally to a much lesser extent. In order to understand the causes of this prolonged contamination, a comparative study on PSP decontamination between sweet clam and cockle was conducted from November 2001 until June 2002. PSP toxicity was analysed by automated pre-column oxidation (Prechromatographic oxidation and LC-FD) in several organs of both species, namely digestive gland, foot, gill, mantle, muscle and siphon for sweet clams. The results showed that cockle sequester PSP toxins preferably in non-visceral organs (Foot, gill and mantle) contrary to sweet clam that sequester them in visceral tissues (digestive gland). The toxin profile of cockle organs indicated dominance of dcSTX, whereas sweet clam tissues contained especially C-toxins. Substantial differences in toxin profile between cockle and sweet clam, from the same area as well as from the composition of PSP toxin producer, Gymnodinium catenatum, confirm the bioconversion of PSP toxins in cockle.     

Paralytic shellfish poisoning toxin profile of mussels Perna perna from southern Atlantic coasts of Morocco

During the monitoring programme of harmful algal blooms established along the south Atlantic coast of Morocco, a bimonthly determination of harmful algae and phycotoxins analysis in Perna perna was carried out from May 2003 to December 2004. Results of mouse bioassay (in organs and whole flesh) showed a seasonal evolution of paralytic shellfish poisoning (PSP) toxin. The mussel's contamination was associated with the occurrence in water of Alexandrium minutum.The PSP toxin profile obtained with high-performance liquid chromatography (HPLC/FD) revealed the dominance of gonyautoxins GTX2 and GTX3 and a minority of GTX1, GTX4 and saxitoxin (STX). This profile explains that the toxicity was mainly associated with A. minutum.     

Determination of paralytic shellfish poisoning toxins in Norwegian shellfish by liquid chromatography with fluorescence and tandem mass spectrometry detection.

A novel extraction and clean-up method has been developed for the determination of paralytic shellfish poisoning (PSP) toxins in shellfish samples. Raw shellfish material was extracted with an acidic acetonitrile/water (80:20, v/v) solution, whilst being homogenised. During the homogenisation the sample extraction solution was cooled with ice water. Subsequently, the extract was frozen at -20 degrees C for at least 4h. During freezing, two layers were formed, only the lower predominantly aqueous layer was used for the determination. The final extract solution was cleaned-up using a combination of Oasis HLB and Carbograph activated carbon SPE columns. The developed extraction and clean-up methods combined with gradient elution liquid chromatography (LC)-mass spectrometry/mass spectrometry (MS/MS) has resulted in a method which can determine the analogs GTX 1-5, C1-2, DcGTX 2-3, DcSTX, Neo, STX in a single analysis with an overall detection limit of 313mug STXdiHCL-eq./kg shellfish meat. The use of the developed extraction method with post-column high performance liquid chromatography (HPLC) with fluorescence detection (FLD) method provided an overall limit of detection of 89mug STXdiHCL-eq./kg shellfish meat for the same toxins. Both post-column HPLC-FLD and LC-MS/MS was used to investigate the Norwegian PSP toxin profile. It was found that the PSP toxins could be detected in shellfish samples from the Norwegian coastline for 10 months of the year, from March till December. The toxin profile consisted mainly of the carbamate toxins, GTX 1-4, Neo and STX, in terms of both concentrations and contribution to the overall toxicity. In addition, several of the n-sulfo-carbamoyl toxins were either detected in the samples at relatively low concentrations or their presence in the samples were indicated but could not be confirmed by the post-column HPLC-FLD and LC-MS/MS analyses.     

Comparative analysis of pre- and post-column oxidation methods for detection of paralytic shellfish toxins

Paralytic shellfish poisoning (PSP) toxins are highly toxic natural compounds produced by dinoflagellates commonly present in marine phytoplankton. Shellfish contaminated with these toxins create significant public health threat and economic losses to the shellfish industry. For this reason, several methods of high performance liquid chromatography (HPLC) with fluorescence detection have been developed in order to gain better knowledge of toxins profiles in shellfish and dinoflagellates samples. These methods have been subjected to continuous modifications to improve and shorten the run time of analysis in the routine monitoring control. In this paper, different samples are analyzed by pre- and post- column HPLC methods to compare toxin profiles. All PSP toxins were individually identified and quantified within the post-column oxidation method. However, although the pre-column oxidation method is significantly more sensitive and detects lower toxin levels, it provides a total amount of toxins that co-elute together, as GTX2 and 3, GTX1 and 4 and dcGTX2 and dcGTX3. The results obtained by both HPLC methods showed similar toxin concentration (expressed in μg/mL) in mussel samples, however when dinoflagellates samples were analyzed the toxin profile and concentration were different. In summary, the post-column oxidation method is accurate to determine the amount of each individual PSP toxin and to know the real toxic profile of non-transformed samples. In addition, this method is easy and faster to screen a large number of samples. The pre-column HPLC method is useful when mussel samples are analyzed even though the time required to prepare the samples is longer.     

Studies in the use of magnetic microspheres for immunoaffinity extraction of paralytic shellfish poisoning toxins from shellfish

Paralytic shellfish poisoning (PSP) is a potentially fatal human health condition caused by the consumption of shellfish containing high levels of PSP toxins. Toxin extraction from shellfish and from algal cultures for use as standards and analysis by alternative analytical monitoring methods to the mouse bioassay is extensive and laborious. This study investigated whether a selected MAb antibody could be coupled to a novel form of magnetic microsphere (hollow glass magnetic microspheres, brand name Ferrospheres-N) and whether these coated microspheres could be utilized in the extraction of low concentrations of the PSP toxin, STX, from potential extraction buffers and spiked mussel extracts. The feasibility of utilizing a mass of 25 mg of Ferrospheres-N, as a simple extraction procedure for STX from spiked sodium acetate buffer, spiked PBS buffer and spiked mussel extracts was determined. The effects of a range of toxin concentrations (20-300 ng/mL), incubation times and temperature on the capability of the immuno-capture of the STX from the spiked mussel extracts were investigated. Finally, the coated microspheres were tested to determine their efficiency at extracting PSP toxins from naturally contaminated mussel samples. Toxin recovery after each experiment was determined by HPLC analysis. This study on using a highly novel immunoaffinity based extraction procedure, using STX as a model, has indicated that it could be a convenient alternative to conventional extraction procedures used in toxin purification prior to sample analysis.     

Comparative determination of paralytic shellfish toxins (PSTs) using five different toxin detection methods in shellfish species collected in the Aleutian Islands, Alaska

Paralytic shellfish poisoning (PSP), a human illness caused by the ingestion of shellfish contaminated with paralytic shellfish toxins (PSTs), has been reported in Alaska for decades. These poisoning incidents have resulted in losses to local economies due to shellfish harvest closures. Thus the development of an effective biotoxin monitoring program designed specifically for the remote regions of Alaska would provide protection for public health and allow for a viable shellfish industry. The present study provides data useful for the development of an effective toxin screening protocol by comparing PST levels quantified in shellfish by many of the currently available PST detection techniques. Seven bivalve species were collected along beaches of the Aleutian Islands from June 2006 to September 2007. The concentration of PSTs was quantified and compared using five different analytical methods: the mouse bioassay, high performance liquid chromatography (HPLC), receptor-binding assay, the commercially available Jellett Rapid PSP Test strips, and an enzyme linked immunosorbent assay technique. The Association of Official Analytical Chemists (AOAC)-approved HPLC method proved to be valuable for characterizing the suite of individual PSTs in each species for research purposes, but was not considered practical for rapid toxin screening in remote Alaskan regions due to its time-consuming nature and requirement of expensive equipment and considerable expertise. In the present study, Jellett test strips were shown to be an effective tool for rapid screening, however due to the high percentage of false positives, subsequent validation via AOAC-approved methods would be required to prevent unnecessary closures.     

Accumulation and depuration of paralytic shellfish poisoning toxins by laboratory cultured purple clam Hiatula diphos Linnaeus.

Purple clams (Hiatula diphos Linnaeus) accumulate paralytic shellfish poisoning (PSP) toxins produced by a toxic strain of the dinoflagellate Alexandrium minutum Halim in a laboratory study. The maximal toxicity of PSP toxins attained 31.3m MU/g after 20 days exposure. The toxin profile of H. diphos was similar to that reported for A. minutum at the end of the exposure period; and GTX1 was dominant. GTX congeners were found in muscle on day 16 and day 20, these substances could be detected during the depuration period as well. GTX1 was detected in the siphon only on day 32. The results show that H. diphos accumulates PSP toxins according to the amount and toxin profile of ingested A. minutum.     

Determination of paralytic shellfish toxins in Portuguese shellfish by automated pre-column oxidation.

Automated pre-column oxidation (the method of Lawrence) was implemented on a routine basis since the end of 1996 to study paralytic shellfish poisoning (PSP) toxins in Portuguese shellfish. Liquid chromatography confirmed the presence of PSP toxins when the known toxic algae were present: Gymnodinium catenatum and/or Alexandrium cf. lusitanicum. On the other side, it has eliminated PSP toxins as a possible recurrent contaminant in oysters from Sado estuary. These oysters were already known to contain high levels of some metals (mainly zinc, copper and cadmium) due to their location in a contaminated area and their particular physiology prone to accumulate metals. The presence of PSP toxins in Scrobicularia plana from Mondego estuary and Tellina crassa from the northern coast, during the absence of the above toxic microalgae in the water column, was confirmed. Unlike other shellfish, these two genera have the feeding habit of aspirating more sediment than organisms in suspension, and probably ingest from the sediment resting cysts of PSP producing microalgae. This is another route of contamination that may help to explain why after a bloom certain shellfish species maintain toxicity for long periods. The method revealed to have a fast implementation on a daily basis, short analysis time (around 20 min between samples), high sensitivity and robustness, and therefore, it is one of the best HPLC methods for screening a large number of shellfish samples for monitoring purposes.     

Frequent occurrence of paralytic shellfish poisoning toxins as dominant toxins in marine puffer from tropical water.

Considerably high toxicity was detected in marine puffers collected from Masinloc Bay, Philippines. The toxicity was detected in the liver, intestine, muscle and skin. Noteworthy, the specimens, the muscle of which showed high toxicity, appeared in high frequency, indicating that puffers from this area is not safe for human consumption. These puffer specimens contained paralytic shellfish poisoning (PSP) toxins, often as major toxin components, the profile of which was similar to that of freshwater puffers reported from tropical areas. These results indicate that PSP toxins are common in tropical puffers both from marine and freshwater.     

Occurrence of paralytic shellfish poison (PSP) in the starfish Asterina pectinifera collected from the Kure Bay, Hiroshima Prefecture, Japan.

Assays were made for paralytic toxicity of marine invertebrates inhabiting at the coasts of Hiroshima Bay, where the infestation of bivalves such as cultured oysters with paralytic shellfish poison (PSP) has been occurred. The starfish Asterina pectinifera collected at the estuary of Nikoh River, Hiroshima Bay, was found to contain moderate levels of paralytic toxicity. Its highest toxicities as PSP found on July 30, 1999 were 12.5 MU/g for whole body, 11.0 MU/g for integument tissues and 3.9 MU/g for viscera, respectively. The toxicity of integument was changed from 3.6 to 11.0 MU/g in 1 year. Its paralytic toxin principles were identified as PSP toxins, composing mainly from saxitoxin (STX) group toxins such as carbamoyl-N-hydroxy neosaxitoxin (hyneoSTX), and STX, by HPLC and LC-MS, accounting for over 90 mol%. The PSP toxins contained in the starfish A. pectinifera considered to be transferred from bivalves or detritus living in the same area, which were contaminated with PSP. However, the involved pathway may be different from that of Asterias amurensis which was infested directly through food chain from its food bivalves, for its toxin pattern.     

Non-selective retention of PSP toxins by the mussel Mytilus galloprovincialis fed with the toxic dinoflagellate Alexandrium tamarense.

Mussels, Mytilus galloprovincialis, were contaminated by paralytic shellfish poisoning (PSP) toxins by being fed with the toxic dinoflagellate Alexandrium tamarense. Temporal variations in the toxin content and the profile of mussels during the feeding experiment were monitored by high-performance liquid chromatography (HPLC). The toxin profile of mussels was compared with that of A. tamarense to clarify the mechanism of uptake of toxins in mussels. The prominent toxins in mussels and A. tamarense were N-sulfocarbamoyl toxins (C1,2) and carbamate toxins, gonyautoxin-1,4 (GTX1,4). The toxin profiles of both mussels and A. tamarense were almost constant throughout the experimental period. There were no remarkable differences in the toxin proportion between mussel and A. tamarense. These results indicate that mussels do not selectively accumulate particular toxins.     

Transfer and metabolism of paralytic shellfish poisoning from scallop (Chlamys nobilis) to spiny lobster (Panulirus stimpsoni)

The transfer and transformation of paralytic shellfish poisoning (PSP) from scallop Chlamys nobilis to spiny lobster Panulirus stimpsoni were investigated in the present study. The results demonstrate that transfer and transformation of PSP toxins occurred when Panulirus stimpsoni were fed with toxic viscera of Chlamys nobilis, but depurated with non-toxic squids. Additionally, only the lobster hepatopancreas were found to contain PSP, and the toxin profiles were the same with those in the viscera of the scallop, including carbamate toxins (GTX₁₋₃), N-sulfocarbamoyl toxins (C₁₊₂ and B₁) and decarbamoyl toxins (dcGTX₂₊₃). Unlike the lobster, the scallop contained more than β toxins. After being fed with toxic Chlamys nobili for 6 d, Panulirus stimpsoni selectively accumulated N-sulfocarbamoyl toxins with low toxicity. However, when they were depurated with non-toxic squid, N-sulfocarbamoyl toxins tended to transform into carbamate toxins with higher toxicity. The concentration of dcGTX₂₊₃ in Panulirus stimpsoni decreased significantly and wasn’t detectable after depuration for 6 d, which was likely due to their initial low accumulation of toxins. These results reveal that PSP could be transferred and transformed in Crustaceans along the given food chain under the conditions of laboratory, but there are many questions remained to be solved, and the further studies should be carried out.     

Surveillance of algal toxins in shellfish from Scottish waters.

Shellfish samples were collected from coastal and offshore aquaculture sites and harvesting areas in Scottish waters between March 2003 and September 2004. Samples were analysed for the presence of algal toxins using traditional mouse bioassays for the detection of paralytic shellfish poisoning (PSP) toxins and diarrhetic shellfish poisoning (DSP) toxins; immuno-lateral flow chromatography for the detection of PSP toxins in the form of the Jellett Rapid Test; high-performance liquid chromatography (HPLC) with UV diode-array for the detection of amnesic shellfish poisoning (ASP) toxins; and liquid chromatography with mass spectrometry (LC-MS) for the detection of multiple lipophilic shellfish toxins (LSTs) including pectenotoxins (PTXs), yessotoxins (YTXs), azaspiracids (AZAs) and toxins from the 'traditional' DSP toxin group, okadaic acid (OA) and dinophysistoxins (DTXs). In order to investigate the presence of OA esters, alkaline hydrolysis was performed. All toxin groups were detected with a geographically widespread distribution. ASP toxins were the most prevalent occurring in 69% of samples. Using the PSP mouse bioassay, PSP toxins were detected in 5% of shellfish samples from coastal waters around the islands and the east coast. The Jellett Rapid Test for PSP toxins revealed a wider distribution (24% of samples) including the west coast of Scotland. Toxins from the 'traditional' DSP toxin group (OA/DTXs) and/or other LST groups (PTXs, YTXs and AZAs) were detected by LC-MS in 63% of the shellfish analysed. PSP, ASP toxins and LSTs occurred concurrently in a limited sample set, highlighting the importance of using methods capable of detecting multiple algal toxin groups in Scottish shellfish monitoring programmes.     

A rapid and sensitive assay for paralytic shellfish poison (PSP) toxins using mouse brain synaptoneurosomes.

A membrane potential assay using mouse brain synaptoneurosomes was evaluated for the determination of paralytic shellfish poison (PSP) toxin content of mussels and other bivalve species important to the shellfish industry. The assay relies on the ability of PSP toxins to block veratridine-induced depolarization of synaptoneurosomes. Changes in the membrane potential of synaptoneurosomes were monitored using the voltage-sensitive fluorescent probe rhodamine 6G. Standard saxitoxin was found to be a potent inhibitor of the membrane depolarizing effects of the sodium channel activator veratridine (I(50) ca. 4 nM). Likewise, shellfish extracts containing PSP toxins inhibited veratridine-induced depolarization. Neither saxitoxin or shellfish extracts had any discernible effect on the resting membrane potential of synaptoneurosomes. When synaptoneurosomal results for extracts of mussels (n=120) and other shellfish (n=29) were correlated with official mouse toxicity assay data there was very good agreement (r(2)=0.84 and 0.86, respectively), indicating that the in vitro assay has utility for a variety of commercially relevant shellfish species. Our investigation suggests that the mouse synaptoneurosome assay is of similar sensitivity to the official CD1 mouse toxicity assay. The synaptoneurosome fraction can be prepared quickly (approx. 40 min) and an individual assay takes less than 7 min. Since 20 such assays can be performed using material from a single CD1 mouse brain, there is considerable opportunity for reducing the number of animals required in conventional PSP monitoring while retaining the same animal system.     

Paralytic shellfish poisoning: post-mortem analysis of tissue and body fluid samples from human victims in the Patagonia fjords.

In July 5, 2002 fishermen working in harvesting sea urchin (Loxechinus albus) in the Patagonia Chilean fjords were intoxicated by consumption of filter-feeder bivalve Aulacomya ater. After the ingestion of 7-9 ribbed mussel, two fishermen died 3-4 h after shellfish consumption. The forensic examination in both victims did not show pathological abnormalities with the exception of the lungs conditions, crackling to the touch, pulmonary congestion and edema. The toxic mussel sample showed a toxicity measured by mouse bioassay of 8575 microg of STX (saxitoxin) equivalent by 100 g of shellfish meat. Using post-column derivatization HPLC method with fluorescent on line detection was possible to measure mass amount of each paralytic shellfish poisoning (PSP) toxin yielding individual toxin concentrations. These PSP toxins were identified in the gastric content, body fluids (urine, bile and cerebrospinal fluid) and tissue samples (liver, kidney, lung, stomach, spleen, heart, brain, adrenal glands, pancreas and thyroids glands). The toxin profiles of each body fluid and tissue samples and the amount of each PSP toxin detected are reported. The PSP toxins found in the gastric content, were STX and the gonyautoxins (GTX4, GTX1, GTX5, GTX3 and GTX2) which showed to be the major amount of PSP toxins found in the victims biological samples. The PSP toxin composition in urine and bile showed as major PSP toxins neoSaxitoxin (neoSTX) and GTX4/GTX1 epimers, both STX analogues with an hydroxyl group (-OH) in the N(1) of the tetrahydropurine nucleus. The neoSTX was not present in the gastric content sample, indicating that the oxidation of N(1) in the STX tetrahydropurine nucleus resulted neoSTX, in a similar way that GTX3/GTX2 epimers were transformed in GTX4/GTX1 epimers. Beside this metabolic transformation, also the hydrolysis of carbamoyl group from STX to form its decarbomoyl analogue decarbamoylsaxitoxin was detected in liver, kidney and lung. These two findings show that PSP toxins went under metabolic transformation during the 3-4 h of human intoxication period, in which PSP toxins showed enzymatic oxidation of N(1) in the tetrahydropurine nucleus, producing neoSTX and GTX4/GTX1 epimers starting from STX and GTX3/GTX2 epimers, respectively. This study conclude, that PSP toxins are metabolically transformed by humans and that they are removed from the body by excretion in the urine and feces like any other xenobiotic compound.     

Accumulation and depuration of paralytic shellfish poisoning toxins by purple clam Hiatula rostrata Lighttoot.

Purple clams (Hiatula rostrata Lighttoot) accumulated paralytic shellfish poisoning (PSP) toxins produced by a toxic strain of the dinoflagellate Alexandrium minutum Halim for subsequent study of toxin distribution during depuration (detoxification by a nontoxic microalgal diet or starvation). The results confirm the data in the literature concerning the high toxicity of the digestive gland, and the depuration efficiency between feed with nontoxic microalgae and starvation is similar. The toxin profile of the purple clams was similar with that of Alexandrium minutum at the end of the exposure period; GTX4 and GTX1 were dominant. However, at the end of the depuration period, GTX3 and GTX2 were dominant. The non-visceral tissues were toxic after feeding with toxic algae. The toxicity was low and the profile were also similar with that of the toxic algae. No PSP toxins other than GTX-1, 2, 3 and 4 were detected in the experimental period.     

Cross-reacting antigens in the butter clam (Saxidoma giganteus) and their relationship to total paralytic shellfish poison toxicity.

A specific protein with an apparent mol. wt of 23,000 was identified in foot homogenate derived from paralytic shellfish poisoning (PSP) contaminated butter clams and was found to cross-react with crab-saxitoxin-induced protein (SIP) antiserum. Antiserum, once cross-absorbed against non-toxic shellfish material, was incubated with tissue homogenate derived from 52 butter clams with varying total PSP toxicities in a prototype ELISA. A significant (r = 0.83; P less than 0.001) correlation existed between soluble clam antigen content in foot homogenate and total PSP toxicity; the latter measured by the mouse lethality bioassay. From the ELISA results, a soluble antigen threshold of 0.1% total protein was successfully used to distinguish between PSP toxic and non-toxic butter clams. It is proposed that this type of screening assay could be used in conjunction with the standard mouse bioassay to increase PSP monitoring and potentially reduce unnecessary animal testing.