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Effect of okadaic acid on glucose regulation.

Okadaic acid is the main toxin responsible for the natural phenomena known as diarrheic shellfish poisoning (DSP). This toxin is a tumor promoter C38 polyether fatty acid that contains acidic and hydrophobic moieties and is cyclic. Okadaic acid is a potent inhibitor of important classes of protein serine/threonine phosphatases such as protein phosphatase 1 and 2A. The toxin binds in a hydrophobic groove adjacent to the active site of the protein phosphatases and interacts with basic residues within the active site. Therefore okadaic acid causes increases in phosphorylation of proteins that affect a diverse array of cellular processes. For instance, this toxin modulates metabolic parameters in intact cells. In this sense it stimulates lipolysis, and inhibits fatty acid synthesis in adipocytes however increases glucose output and gluconeogenesis in hepatocytes. Additionally, okadaic acid reaches cytotoxic concentrations in the intestinal tissues in accordance with the diarrhea. Recent studies suggested that toxic effects of okadaic acid might be related to modification of nutrients, ionic and water absorption across the small intestine presumably by altering the transporter system. The subject of this review is limited to the effect of okadaic acid on glucose regulation and its cellular as well as clinical implications.

Animals↗

Potential impacts of climate change on infectious diseases in the Arctic.

Climate change could cause changes in the incidence of infectious diseases in Arctic regions. Higher ambient temperatures in the Arctic may result in an increase in some temperature sensitive foodborne diseases such as gastroenteritis, paralytic shellfish poisoning and botulism. An increase in mean temperature may also influence the incidence of infectious diseases of animals that are spread to humans (zoonoses) by changing the population and range of animal hosts and insect vectors. An increase in flooding events may result in outbreaks of waterborne infection, such as Giardia lamblia or Cryptospordium parvum. A change in rodent and fox populations may result in an increase in rabies or echinococcosis. Temperature and humidity influence the distribution and density of many arthropod vectors which in turn may influence the incidence and northern range of vectorborne diseases such as West Nile virus. Recommendations include: the strenghtening of public health systems, disease surveillance coordinated with climate monitoring, and research into the detection, prevention, control and treatment of temperature-sensitive infectious diseases.

Animals↗

Determination of domoic acid in Japanese mussels by enzyme immunoassay.

Ten samples of commercial blue mussels (Mytilus edulis) from Japan were analyzed for domoic acid by an indirect competitive enzyme immunoassay (idc-EIA) based on an anti-domoic acid monoclonal antibody. Domoic acid was found in all samples at low concentrations (0.11-1.81 ng/g mussel tissue). The presence of domoic acid was confirmed by liquid chromatography coupled with immunoaffinity chromatography using an anti-domoic acid monoclonal antibody as ligand. To our knowledge, this is the first reported detection of domoic acid, a causative agent of amnesic shellfish poisoning, in Japanese mussels.

Animals↗

Feasibility of reduction in use of the mouse bioassay: presence/absence screening for saxitoxin in frozen acidified mussel and oyster extracts from the coast of California with in vitro methods.

In the United States, the detection of paralytic shellfish poisoning (PSP) for regulatory purposes relies on the mouse bioassay (MBA). Using a saxitoxin presence/absence test could reduce animal usage significantly. Three in vitro methods, the RIDASCREEN Saxitoxin kit, MIST Alert, and a 5 h neuroblastoma assay, were evaluated in parallel with the MBA using 106 twice-frozen, acidified extracts from California-grown mussel and oyster tissues. For each assay, a cutoff point was established whereby data below or equal to that point were scored as negative and were assigned a score of zero. Data above the cutoff were considered positive and assigned a score of one. Pearson correlation coefficients were generated. The RIDASCREEN, MIST Alert, and neuroblastoma bioassay correlated to the MBA at 0.849, 0.853, and 0.832 when used for presence/absence detection. These data suggest that a reduction in MBA usage could be achieved in the surveillance of California-grown mussels and oysters for PSP-associated toxins. Correlation data between the in vitro assays, cost comparisons, and the potential for false negatives and false positives were examined. Implications of these methodologies in protecting public health are discussed.

Animals↗

Committee on Natural Toxins and Food Allergens. Marine and freshwater toxins.

There have been major developments this past year in the Marine and Freshwater Toxins topic area (formerly Phycotoxins). These include AOAC approval and inauguration of a new AOAC Presidential Task Force on Marine and Freshwater Toxins to accelerate methods validation, and the appointment of several new Topic Advisors. A joint FAO/IOC/WHO group addressing biotoxins in molluscan bivalves is also relevant to this report and to the new Task Force. The AOAC Presidential Task Force on Marine and Freshwater Toxins is an international group that, in late November 2004, consisted of 90 world experts and stakeholders. Chaired by this General Referee, the group establishes methods priorities based on analytical methods criteria, determines fitness for purpose, identifies and reviews available methodologies, recommends methodologies for validation, and identifies complementary analytical tools. Once appropriate analytical methodology has been identified or developed, the Task Force is able to identify financial and technical resources necessary to validate the methods. The first two formal meetings of the Task Force were held in Bethesda, MD, on May 19, 2004 and in St. Louis, MO, on September 22, 2004. These meetings were held in conjunction with the XI International IUPAC Symposium on Mycotoxins and Phycotoxins and the 118th AOAC INTERNATIONAL Annual Meeting and Exposition, respectively. The Bethesda meeting served to introduce members of the group to the AOAC Community/Task Force model and to discuss objectives, concerns, general workings, and communications. The meeting concluded on an encouraging note, with a commitment from AOAC to help provide financial resources for the review of nonproprietary methods deemed high priority by the Task Force. This development was seen as an important step toward reaching methods validation objectives. The terms of reference for the Task Force were approved by the AOAC Board of Directors in late June, 2004. They described the Task Force membership as composed of voting and nonvoting members, with the voting members consisting of 13 members (12 plus the Chair). Voting members comprise of a balance of government regulators, academics, and industry members. No single agency has more than 2 voting members. Task Force members serve as experts in the field and agree to identify other experts; recommend individuals who can serve on the Task Force and as Chair; develop and prioritize a list of marine and freshwater toxins that need validated methods; assist in identifying existing methods for validation through AOAC validation programs; and recommend to the AOAC INTERNATIONAL Board of Directors policies and procedures necessary to accomplish the mission of the Task Force. They endeavor to actively support the work of the Task Force through garnering of sources of funding (except where prohibited by employer); identifying potential participating laboratories, sample identification and acquisition; and increasing program awareness among stakeholders. They assist AOAC in the identification of study directors and in the development of quality measurement tools by participating in the validation of methods and by identifying venues for members of the Task Group or the community to gather and assist with meeting content. Prior to the September 2004, AOAC Annual Meeting, the Task Force approved a set of Analytical Methods Selection Criteria, which are critical to the mission of the Task Force. They can be found, along with the Terms of Reference, roster of members, and other information, on the Task Force Web site at http://www.aoac.org/marine toxins/task_force.htm. The September 22, 2004 Task Force meeting in St. Louis included discussion of 2 interlaboratory studies, a proprietary kit for domoic acid by enzyme-linked immunosorbent assay (ELISA; Biosense Labs AS, Bergen, Norway) and also a nonproprietary liquid chromatography (LC) method for paralytic shellfish poisoning (PSP) toxins by precolumn oxidation (James F. Lawrence, Health Canada). These 2 methods were recommended by the Task Force for review by AOAC in September 2004. The group also discussed future priority directions, aspects of interlaboratory studies and official methods of analysis, other methods validation issues, future meetings, and funding. In addition to the Task Force meeting, 2 subgroup meetings were held. One subgroup addressed strategies to replace the mouse bioassay for brevetoxins with alternative modern methods based on ELISA or LC/mass spectrometry (MS). Brevetoxin metabolites, toxicity issues, and extraction conditions as well as future field studies were addressed in detail. The receptor binding assay (RBA)/saxitoxins subgroup addressed several aspects of the methodology, radiolabeled saxitoxin, and comparisons of mouse bioassay and RBA response. Both subgroups were productive and were seen as very useful by the participants. Task Force attendees generally agreed that subgroups are the most effective means of progressing towards validation of new methods and of ensuring thorough discussions of methods under consideration. By the time of their next meeting (April 2005) at the "Marine and Freshwater Toxins Analysis: 1st Joint Symposium and AOAC Task Force Meeting" in Baiona, Spain, the Task Force will have several well developed new subgroups in the areas of okadaic acid and dinophysis toxins, yessotoxins, domoic acids, and ciguatoxins. Some of the subgroups will hold face-to-face meetings in Spain and others will meet at future symposia or joint meetings. It is likely that training sessions will be associated with multiple Task Force meetings planned for 2005. Details on these meetings can be found on the Task Force Web site. Although the Task Force has experienced rapid growth, the addition of new members to the group, especially industry and government stakeholders, is encouraged. Task Force member Michael Quilliam, NRC Canada, provided the information given below on a joint CODEX group of special relevance to the new Task Force. This group met in late September 2004. For more information, see http://www.who.int/foodsafety/chem/meetings/biotoxin/en/.

Advisory Committees↗

Final report and recommendations from the National Notifiable Diseases Working Group.

UNLABELLED: The National Notifiable Diseases Working Group performed a ranking of 48 communicable diseases to assist with determining priorities for national surveillance. The WG offers six specific recommendations relating to the addition or deletion of communicable diseases from the list of nationally notifiable diseases. KEY RECOMMENDATIONS: 1. Clostridium difficile-associated diarrhea, paralytic shellfish poisoning, and invasive listeriosis are recommended for addition to the Nationally Notifiable Disease List. 2. New proposals for diseases not recommended for national notification include hepatitis E, HTLV-1, and HTLV-2. 3. Dengue virus infection should be deleted from the Nationally Notifiable Disease List. (Dengue hemorrhagic fever should continue to be reported.) 4. Classic Creutzfeld-Jacob disease ranked well below the threshold for inclusion; however, it should be retained on the list because surveillance of classic CJD is key to effective surveillance of new variant disease and because consulted provinces and territories strongly favoured its retention. 5. Several diseases that ranked near the borderline for reporting should be retained on the list as follows: a. a. brucellosis, because although it does not rank highly on the basis of its endemic pattern it is a category B biological warfare agent; b. laboratory-confirmed influenza because it forms part of a functional and working surveillance approach; c group B streptococcal infection because alternative hospital-based surveillance systems do not yet capture a significant proportion fo cases. 6. Transfusion-transmitted infections were unrankable by the current system. 7. The approach to reporting dengue hemorrhagic fever, plague, West Nile virus infections, and other viral hemorrhagic fevers should be made congruent with the approach for other notifiable diseases (see Discussion). 8. The feasibility and utility of national reporting for communicable diseases that have very high incidence and low severity (e.g. chickenpox, norovirus) should be carefully considered, especially when it leads to little or no case-by-case data reported by health authorities. The WG recommends that these diseases be placed under surveillance but that case-by-case reporting not be required at the national level. Alternative approaches to tracking trends could include enumerating outbreaks in the case of norovirus or the use of sentinel surveillance, laboratory surveillance, or physician billing events in the case of varicella.

Biological Warfare↗

Growth physiology of red-tide microorganisms.

The occurrence of red tides, which often cause mass mortality of marine animals and/or shellfish poisoning in humans, is increasing in the estuaries and coastal zones of the world. Species of dinoflagellates are most commonly responsible, though other flagellates, and even ciliates, can also cause red tides. In recent years, understanding of the growth physiology of red-tide microalgae has grown. Each causative organism has a species-specific preference and requirement for temperature, salinity, pH, the basic nutrients, and growth factors, and the toxin formation is affected by these environmental factors.

Animals↗

Global climate change and emerging infectious diseases.

Climatic factors influence the emergence and reemergence of infectious diseases, in addition to multiple human, biological, and ecological determinants. Climatologists have identified upward trends in global temperatures and now estimate an unprecedented rise of 2.0 degrees C by the year 2100. Of major concern is that these changes can affect the introduction and dissemination of many serious infectious diseases. The incidence of mosquito-borne diseases, including malaria, dengue, and viral encephalitides, are among those diseases most sensitive to climate. Climate change would directly affect disease transmission by shifting the vector's geographic range and increasing reproductive and biting rates and by shortening the pathogen incubation period. Climate-related increases in sea surface temperature and sea level can lead to higher incidence of water-borne infectious and toxin-related illnesses, such as cholera and shellfish poisoning. Human migration and damage to health infrastructures from the projected increase in climate variability could indirectly contribute to disease transmission. Human susceptibility to infections might be further compounded by malnutrition due to climate stress on agriculture and potential alterations in the human immune system caused by increased flux of ultraviolet radiation. Analyzing the role of climate in the emergence of human infectious diseases will require interdisciplinary cooperation among physicians, climatologists, biologists, and social scientists. Increased disease surveillance, integrated modeling, and use of geographically based data systems will afford more anticipatory measures by the medical community. Understanding the linkages between climatological and ecological change as determinants of disease emergence and redistribution will ultimately help optimize preventive strategies.

Animals↗

Disorders of neuromuscular transmission due to natural environmental toxins.

A variety of natural toxins of animal, plant, and bacterial origin are capable of causing disorders of neuromuscular transmission. Animal toxins include venomous snakes and arthropods, venoms of certain marine creatures, skin secretions of dart-poison frogs, and poisonous fish, shellfish, and crabs. There are plant poisons such as curare, and bacterial poisons such as botulinum toxin. These act at single or multiple sites of the neuromuscular apparatus interfering with voltage-gated ion channels, acetylcholine release, depolarization of the postsynaptic membrane, or generation and spread of the muscle action potential. The specific actions of these toxins are being widely exploited in the study of neuromuscular physiology and pathology. Some toxins have proved to be valuable pharmaceutical agents. Poisoning by natural neurotoxins is an important public health hazard in many parts of the world, particularly in the tropics. Poisoning may occur by a bite or a sting of a venomous animal, or by the ingestion of poisonous fish, shellfish or other marine delicacies. Contaminated food is a vehicle for poisons such as botulinum toxin. Clinically, a cardinal feature in the symptomatology is muscle paralysis with a distribution characteristic of myasthenia gravis, affecting muscles innervated by cranial nerves, neck flexors, proximal limb muscles, and respiratory muscles. Respiratory paralysis may end fatally. This paper reviews from the clinical and pathophysiologic viewpoints, naturally occurring environmental neurotoxins acting at the neuromuscular junction.

Animals↗

Azaspiracid poisoning, the food-borne illness associated with shellfish consumption.

Azaspiracid poisoning (AZP) is a recently discovered toxic syndrome that was identified following severe gastrointestinal illness from the consumption of contaminated mussels (Mytilus edulis). The implicated toxins, azaspiracids, are polyethers with unprecedented structural features. Studies toward total toxin synthesis revealed that the initial published structures were incorrect and they have now been revised. These toxins accumulate in bivalve molluscs that feed on toxic microalgae of the genus Protoperidinium, previously considered to be toxicologically benign. Although first identified in shellfish from Ireland, azaspiracid contamination of several types of bivalve shellfish species has now been confirmed throughout the western coastline of Europe. Toxicological studies have indicated that azaspiracids can induce widespread organ damage in mice and that they are probably more dangerous than previously known classes of shellfish toxins. The exclusive reliance on live animal bioassays to monitor azaspiracids in shellfish failed to prevent human intoxications. This was a consequence of poor sensitivity of the assay and the fact that azaspiracids are not exclusively found in the shellfish digestive glands used for toxin testing. The strict regulatory control of azaspiracids in shellfish now requires frequent testing of shellfish using highly specific and sensitive methods involving liquid chromatography-mass spectrometry.

Animals↗

Presence of saxitoxin in toxic extracts from Gonyaulax polyedra.

A "red tide" bloom of Gonyaulax polyedra occurred in Italy in Autumn, 1988. Algal concentrated extracts and undiluted water samples from the bloom were tested both with the Microtox system and a mouse bioassay, revealing the presence of paralytic shellfish poison-like neurotoxins. Saxitoxin levels evaluated on the basis of toxicological and instrumental analysis showed discrepancies. Other toxins could be present in addition to paralytic shellfish poison.

Animals↗

Liquid chromatography with electrospray ion trap mass spectrometry for the determination of five azaspiracids in shellfish.

Azaspiracid poisoning (AZP) is a new human toxic syndrome that is caused by the consumption of shellfish that have been feeding on harmful marine microalgae. A liquid chromatography-mass spectrometry (LC-MS) method has been developed for the determination of the three most prevalent toxins, azaspiracid (AZA1), 8-methylazaspiracid (AZA2) and 22-demethylazaspiracid (AZA3) as well as the isomeric hydroxylated analogues, AZA4 and AZA5. Separation of five azaspiracids was achieved on a C18 column (Luna-2, 150 x 2 mm, 5 microm) with isocratic elution using acetonitrile-water containing trifluoroacetic acid and ammonium acetate as eluent modifiers. Using an electrospray ionisation (ESI) source with an ion-trap mass spectrometer, the spectra showed the protonated molecules, [M+H]+, with most major product ions due to the sequential loss of two water molecules. A characteristic fragmentation pathway that was observed in each azaspiracid was due to the cleavage of the A-ring at C9-C10 for each toxin. It was possible to select unique ion combinations to distinguish between the isomeric azaspiracids, AZA4 and AZA5. Highly sensitive LC-MS3 analytical methods were compared and the detection limits were 5-40 pg on-column. Linear calibrations were obtained for AZA1 in shellfish in the range 0.05-1.00 microg/ml (r2 = 0.9974) and good reproducibility was observed with a relative standard deviation (%RSD) of 1.8 for 0.9 microg AZAI/ml (n=5). The %RSD values for the minor toxins, AZA4 and AZA5, using LC-MS3 (A-ring fragmentation) were 12.3 and 8.1 (0.02 microg/ml; n=7), respectively. The selectivity of toxin determination was enhanced using LC-MS-MS with high energy WideBand activation.

Animals↗

First evidence of an extensive northern European distribution of azaspiracid poisoning (AZP) toxins in shellfish.

Azaspiracids have recently been identified as the toxins responsible for a series of human intoxications in Europe since 1995, following the consumption of cultured mussels (Mytilus edulis) from the west coast of Ireland. Liquid chromatography-mass spectrometric (LC-MS) methods have been applied in the study reported here to investigate the new human toxic syndrome, azaspiracid poisoning. Separation of azaspiracid (AZA1) and its analogues, 8-methylazaspiracid (AZA2) and 22-demethylazaspiracid (AZA3), was achieved using reversed-phase LC and coupled, via an electrospray ionisation source, to an ion-trap mass spectrometer. These azaspiracids have now been identified in mussels from Craster (north-east England) and Sognefjord (south-west Norway) using source collision induced dissociation-MS and multiple tandem MS detection. AZA1 was the predominant toxin and toxin profiles were similar to those found in contaminated Irish shellfish. This is the first report of the occurrence of these azaspiracids outside Ireland with the significant implications that these toxins may occur in shellfish throughout northern Europe.

Animals↗

Geographical, temporal, and species variation of the polyether toxins, azaspiracids, in shellfish.

Azaspiracid Poisoning (AZP) is a new toxic syndrome that has caused human intoxications throughout Europe following the consumption of mussels (Mytilus edulis), harvested in Ireland. Shellfish intoxication is a consequence of toxin-bearing microalgae in the shellfish food chain, and these studies demonstrated a wide geographic distribution of toxic mussels along the entire western coastal region of Ireland. The first identification of azaspiracids in other bivalve mollusks including oysters (Crassostrea gigas), scallops (Pecten maximus), clams (Tapes phillipinarium), and cockles (Cardium edule) is reported. Importantly, oysters were the only shellfish that accumulated azaspiracids at levels that were comparable with mussels. The highest levels of total azaspiracids (microg/g) recorded to-date were mussels (4.2), oysters (2.45), scallops (0.40), cockles (0.20), and clams (0.61). An examination of the temporal variation of azaspiracid contamination of mussels in a major shellfish production area revealed that, although maximum toxin levels were recorded during the late summer period, significant intoxications were observed at periods when marine dinoflagellate populations were low. Although human intoxications have so far only been associated with mussel consumption, the discovery of significant azaspiracid accumulation in other bivalve mollusks could pose a threat to human health.

Animals↗