Health Standards for Live Bivalve Molluscs

Food business operators (FBOs) must ensure that live bivalve molluscs (e.g., clams, oysters, cockles, mussels, scallops) placed on the market for human consumption meet the following standards:

They must look, feel, smell and taste like they are fresh and suitable for consumption, including having shells free of dirt, will return to their shape if touched and have normal amounts of intravalvular liquid in them;
They must not contain marine biotoxins in total quantities (measured in the whole body or any part edible separately) that exceed the following limits:

for paralytic shellfish poison (PSP), 800 micrograms per kilogram;
for amnesic shellfish poison (ASP), 20 milligrams of domoic acid per kilogram;
for okadaic acid, dinophysistoxins and pectenotoxins together, 160 micrograms of okadaic acid equivalents per kilogram;
for yessotoxins, 3.75 milligrams of yessotoxin equivalent per kilogram; and
for azaspiracids, 160 micrograms of azaspiracid equivalents per kilogram.

Annex III of Regulation (EC) 2074/2005 (this opens in a new tab) requires that the following analytical methods must be used by the competent authorities to check compliance with the limits listed above (Chapter V (2) of Section VII of Annex III to Regulation (EC) No 853/2004 (this opens in a new tab) and its amendment in Regulation (EU) 786/2013) (this opens in a new tab). In accordance with Article 7(2) and (3) of Council Directive 86/609/EEC (no longer in force) (this opens in a new tab) on the approximation of laws, regulations and administrative provisions of the Member States regarding the protection of animals used for experimental and other scientific purposes elements of replacement, refinement and reduction must be taken into account when biological methods are used.

Paralytic Shellfish Poison (PSP) Detection Method

The paralytic shellfish poison (PSP) content of edible parts of molluscs (the whole body or any part edible separately) must be detected in accordance with the biological testing method or any other internationally recognised method. The so-called Lawrence method may also be used as an alternative method for the detection of those toxins as published in AOAC Official Method 2005.06 (Paralytic Shellfish Poisoning Toxins in Shellfish) (this opens in a new tab).

If the results are challenged, the reference method shall be the biological method.

Amnesic Shellfish Poison (ASP) Detection Method

Lipophilic Toxin Detection Methods - Biological methods

A series of mouse bioassay procedures (a mice bioassay is literally using a mouse to determine how safe a food is), differing in the test portion (hepatopancreas or whole body) and in the solvents used for extraction and purification, may be used for detecting marine toxins as referred to in Chapter V(2)(c), (d) and (e) of Section VII of Annex III, to Regulation (EC) No 853/2004 (okadaic acid, dinophysistoxins and pectenotoxins; yessotoxins and azaspiracids) (this opens in a new tab). Sensitivity and selectivity depend on the choice of solvents used for extraction and purification and this should be taken into account when a decision is made on the method to be used in order to cover the full range of toxins.

A single mouse bioassay involving acetone extraction may be used to detect okadaic acid, dinophysistoxins, pectenotoxins and yessotoxins. This assay may be supplemented, if necessary, with liquid/liquid partition steps with ethyl acetate/water or dichloromethane/water to remove potential interferences. Azaspiracid detection at regulatory levels by means of this procedure shall involve the use of the whole body as the test portion.

Alternative detection methods

A series of methods, such as high-performance liquid chromatography (HPLC) with fluorimetric detection, liquid chromatography (LC), mass spectrometry (MS), immunoassays and functional assays, such as the phosphatase inhibition assay, may be used as alternatives or supplementary to the biological testing methods, provided that either alone or combined they can detect at least the following analogues, that they are not less effective than the biological methods and that their implementation provides an equivalent level of public health protection:

okadaic acid and dinophysistoxins: a hydrolysis step may be required to detect the presence of DTX3;
pectenotoxins: PTX1 and PTX2;
yessotoxins: YTX, 45 OH YTX, homo YTX, and 45 OH homo YTX;
azaspiracids: AZA1, AZA2 and AZA3.

If new analogues of public health significance are discovered, they should be included in the analysis. Standards must be available before chemical analysis is possible. Total toxicity shall be calculated using conversion factors based on the toxicity data available for each toxin. The performance characteristics of these methods shall be defined after validation following an internationally agreed protocol.

Biological methods shall be replaced by alternative detection methods as soon as reference materials for detecting the toxins prescribed in Chapter V of Section VI of Annex III to Regulation (EC) No 853/2004 (this opens in a new tab) are readily available, and the methods have been validated.