A recombinant protein of a Paragonimus westermani egg antigen was produced and tested as an antigen for the serologic diagnosis of P. westermani infection. The P. westermani egg antigen gene contains a single open reading frame of 966 base pairs encoding 322 amino acids from 5' methionine to the 3' stop codon. The predicted amino acid sequence of this egg antigen was 40, 38, and was 35% identical to heat shock proteins from Schistosoma japonicum, Schistosoma mansoni, and Taenia saginata. The distribution this antigen was investigated in adult worms by indirect immunofluorescence assay, and found to be distributed in eggs and uteri. The specificity and sensitivity of the recombinant antigen were assessed by enzyme-linked immunosorbent assay (ELISA) using sera from patients infected with different parasites, which included 41 patients with paragonimiasis, and negative controls. The diagnostic positive and negative predictive absorbance value was 0.24 and the sensitivity of ELISA using the recombinant antigen was 90.2%, and its specificity 100%. Our results suggest that the developed recombinant major egg antigen-based ELISA offers a highly sensitive and specific assay for the diagnosis of paragonimiasis.
The storage mite, Tyrophagus putrescentiae, is one of the important causes of allergic disorders. Fifteen allergenic components were demonstrated in storage mite by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting, but only the group 2 allergen Tyr p 2 has been cloned and characterized. In this study, we attempted to identify and characterize new allergens from T. putrescentiae, which is a dominant species of storage mite in Korea. Expressed sequence tags were analyzed to identify possible storage mite allergens, and the cDNA sequence encoding a protein homologous to fatty acid binding protein, a mite group 13 allergen, was identified and named Tyr p 13. Its deduced amino acid sequence showed 61.1 to 85.3% identity with other mite group 13 allergens. The recombinant protein was expressed in Escherichia coli using a pET 28b vector system, and its allergenicity was investigated by enzyme-linked immunosorbent assay (ELISA). The recombinant allergen was detected in 5 of 78 (6.4%) T. putrescentiae-positive sera tested, and it inhibited 61.9% of immunoglobulin E binding to crude extract at an inhibitor concentration of 10 g/ml by inhibition ELISA using serum from the patient who showed the strongest reaction by ELISA. In this study, a novel allergen was identified in T. putrescentiae. This allergen could be helpful for more-detailed characterizations of storage mite allergy.
Storage mites may cause allergic respiratory diseases in urban areas as well as pose an occupational hazard in rural areas. Characterization of storage mite allergens is important for the development of diagnostic and therapeutic agents against mite-associated allergic disorders. Here we report on the cloning and expression of ␣-tubulin from the storage mite (Tyrophagus putrescentiae). The deduced amino acid sequence of the ␣-tubulin from the storage mite showed as much as 97.3% identity to the ␣-tubulin sequences from other organisms. The highly conserved amino acid sequences of ␣-tubulins across different species of mites may indicate that cross-reactivity for this potential allergen exists. The frequency of immunoglobulin E reactivity of this recombinant protein is 29.3% in sera from storage mite-allergic subjects.
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