Human
angiostrongyliasis results from accidental
infection with Angiostrongylus, an intra-arterial nematode. Angiostrongylus cantonensis
infections result in eosinophilic
meningitis, and A. costaricensis
infections cause
eosinophilic enteritis. Immunological methodologies are critical to the diagnosis of both
infections, since these parasites cannot be isolated from fecal matter and are rarely found in cerebrospinal fluid samples. A. costaricensis and A. cantonensis share common antigenic
epitopes which elicit
antibodies that recognize
proteins present in either species. Detection of
antibodies to a 31-kDa A. cantonensis
protein present in crude adult worm extracts is a sensitive and specific method for immunodiagnosis of cerebral
angiostrongyliasis. The objective of the present work was to isolate and characterize the 31-kDa
proteins using soluble
protein extracts derived from adult female worms using both one- (1DE) and two-dimensional (2DE) gel electrophoresis. Separated
proteins were blotted onto
nitrocellulose and probed using sera from infected and non-infected controls. The 31-kDa band present in 1DE
gels and the 4 spots identified in 2DE
gels were excised and analyzed by electrospray ionization mass spectrometry. Using the highest scores obtained following Mascot analysis, amino acid sequences were obtained that matched four unique
proteins:
tropomyosin, the 14-3-3
phosphoserine-
binding protein, a
protein containing a
nascent polypeptide-associated complex domain, and the putative epsilon subunit of
coatomer protein complex
isoform 2. Oxidative cleavage of diols using
sodium m-
periodate demonstrated that
carbohydrate moieties are essential for the antigenicity of all four spots of the 31-kDa
antigen. In this article we describe the identification of the 31-kDa
antigen, and provide
DNA sequencing of the targets. In conclusion, these data suggest that reactivity to the 31-kDa
proteins may represent antibody recognition of more than one
protein, and
recombinant protein-based assays for cerebral
angiostrongyliasis diagnosis may require eukaryotic expression systems to maintain antigenicity.