title: Quantification of Anisakis simplex allergens in fresh, long-term frozen, and cooked fish muscle.
abstract: Fish-borne parasitic zoonoses such as Anisakiasis were once limited to people living in countries where raw or undercooked fish is traditionally consumed. Nowadays, several factors, such as the growing international markets, the improved transportation systems, the population movements, and the expansion of ethnic ways of cooking in developed countries, have increased the population exposed to these parasites. Improved diagnosis technology and a better knowledge of the symptoms by clinicians have increased the Anisakiasis cases worldwide. Dietary recommendations to Anisakis-sensitized patients include the consumption of frozen or well-cooked fish, but these probably do not defend sensitized patients from allergen exposure. The aim of our work was to develop a sensitive and specific method to detect and quantify Anisakis simplex allergens in fish muscle and its derivatives. Protein extraction was made in saline buffer followed by preparation under acid conditions. A. simplex antigens were detected by IgG immunoblot and quantified by dot blot. The allergenic properties of the extracts were assessed by IgE immunoblotting and basophil activation test. We were able to detect less than 1 ppm of A. simplex antigens, among them the allergen Ani s 4, in fish muscle with no cross-reactions and with a recovery rate of 82.5%. A. simplex antigens were detected in hakes and anchovies but not in sardines, red mullets, or shellfish. We detected A. simplex allergens in cooked hakes and also in hake stock. We proved that A. simplex allergens are preserved in long-term frozen storage (-20 degrees C +/- 2 degrees C for 11 months) of parasitized hakes. Basophil activation tests have proven the capability of the A. simplex-positive fish extracts to induce allergic symptoms.
authors.affiliation: Fundación para la Investigación Biomédica, Hospital Carlos III, Madrid, Spain. anai_rm@yahoo.es
authors.forename: Ana Isabel
authors.initials: AI
authors.lastname: Rodríguez-Mahillo
authors.forename: Miguel
authors.initials: M
authors.lastname: González-Muñoz
authors.forename: Cristina
authors.initials: C
authors.lastname: de las Heras
authors.forename: Margarita
authors.initials: M
authors.lastname: Tejada
authors.forename: Ignacio
authors.initials: I
authors.lastname: Moneo
mesh_terms: D000485:Allergens / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D000818:Animals; D017163:Anisakis / Q000276:immunology* / Q000302:isolation & purification; D015703:Antigens, CD / Q000378:metabolism; D000947:Antigens, Helminth / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D001491:Basophils / Q000276:immunology / Q000378:metabolism; D049872:Bivalvia / Q000469:parasitology; D005396:Fish Products / Q000009:adverse effects / Q000032:analysis / Q000469:parasitology; D005399:Fishes / Q000254:growth & development / Q000469:parasitology*; D005511:Food Handling; D005512:Food Hypersensitivity / Q000097:blood / Q000178:diet therapy / Q000276:immunology / Q000469:parasitology; D005513:Food Inspection / Q000379:methods; D016266:Food Parasitology; D015801:Helminth Proteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D057230:Limit of Detection; D009132:Muscles / Q000737:chemistry / Q000469:parasitology; D033561:Penaeidae / Q000469:parasitology; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D017747:Seafood / Q000009:adverse effects / Q000032:analysis* / Q000469:parasitology*; D012758:Shellfish / Q000009:adverse effects / Q000032:analysis / Q000469:parasitology; D013696:Temperature; D060149:Tetraspanin 30; D014020:Tissue Extracts / Q000009:adverse effects / Q000737:chemistry / Q000276:immunology
chemical_list: D000485:Allergens; C553595:Ani s 3 allergen, Anisakis simplex; D015703:Antigens, CD; D000947:Antigens, Helminth; C554586:CD63 protein, human; D015801:Helminth Proteins; D010980:Platelet Membrane Glycoproteins; D060149:Tetraspanin 30; D014020:Tissue Extracts; D007073:Immunoglobulin E