Characterisation of potential novel allergens in the fish parasite Anisakis simplex

Abstract

The parasitic nematode Anisakis simplex occurs in fish stocks in temperate seas. A. simplex contamination of fish products is unsavoury and a health concern considering human infection with live larvae (anisakiasis) and allergic reactions to anisakid proteins in seafood. Protein extracts of A. simplex produce complex band patterns in gel electrophoresis and IgE-immunostaining. In the present study potential allergens have been characterised using sera from A. simplex-sensitised patients and proteome data obtained by mass spectrometry. A. simplex proteins were homologous to allergens in other nematodes, insects, and shellfish indicating cross-reactivity. Characteristic marker peptides for relevant A. simplex proteins were described.

Keywords: Allergenicity; Anisakis simplex; Cross-reactivity; Immunoblotting; Mass spectrometry; Proteomic analysis.

Fig. 1

(a) IgE-immunoblot with total A. simplex extract using sera from14 Norwegian (N1–N14) and 13 Spanish (S1–S13) patients with sensitivity to A. simplex (s. Table 2). Relevant lanes were extracted from the scanned individual patient immunoblots and presented as a composite gel. M: SeeBluePlus2 molecular weight marker; relative protein molecular weights (kDa) are indicated on the right. (b) Bar Chart showing summarised binding intensities of 27 patients for 22 IgE-binding signals on immunoblot (Fig. 1a). Protein molecular weights (kDa) as determined by GelPro Analyzer^® are given on the ordinate, protein amounts (ng), calculated as described in Section 2, are on the abscissa. The contributions of individual patients to the combined results are marked by different patterns as shown in the legend.

Fig. 2

Coomassie-stained SDS-PAGE of total A. simplex protein extract. Gel bands (No. 1–16) were excised, digested with trypsin, and subjected to mass spectrometric analysis. SeeBlue Plus2 molecular weight marker; relative protein sizes (kDa) are given on the left side of the gel.