Immunoblot analysis of salivary allergens in 10 mosquito species with worldwide distribution and the human IgE responses to these allergens,☆☆,,★★

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Abstract

Background: Most people develop skin reactions to mosquito bites, however, little is known about mosquito salivary allergens and the IgE responses to them. Objectives: We sought to identify these allergens and the specific IgE responses they elicit. Methods: Saliva or salivary gland extracts were prepared from 10 mosquito species, including seven species with worldwide distribution: Aedes (Ae.) aegypti, Ae. vexans, Ae. albopictus, Ae. togoi, Ae. triseriatus, Culex (Cx.) quinquefasciatus, Cx. pipiens, Cx. tarsalis, Anopheles (An.) sinensis, and Culiseta (Cs.) inornata. Proteins from these preparations were separated by sodium dodecylsulfate–polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes, which were immunoblotted by sequential incubations with human serum, monoclonal anti-human IgE, and enzyme-conjugated goat anti-mouse IgG. Salivary allergens were analyzed by using a pooled serum from subjects allergic to mosquitos. Individual IgE responses to each allergen were evaluated in 12 subjects allergic to mosquitos living in Canada, the United States, and China, as well as in five subjects not allergic to mosquito bites. To study species-shared allergens, the membranes were immunoblotted with two rabbit antibodies specific to recombinant mosquito salivary proteins. Results: Three to sixteen salivary allergens with molecular masses ranging from 16 to 95 kd were found in each species. Both species-shared and species-specific allergens were identified by molecular masses, binding to the two rabbit antibodies, and individual IgE responses to species indigenous to and absent from the regions where the subjects lived. Salivary allergens, especially from Ae. aegypti, Ae. vexans, and Ae. albopictus, elicited higher IgE responses in subjects allergic to mosquitos than in nonallergic subjects. Conclusions: Species-shared and species-specific allergens that cause IgE responses in subjects allergic to mosquitos are immunologically identified. Species-shared allergens are the most important for potential use in diagnosis and immunotherapy. (J Allergy Clin Immunol 1998;101:498-505.)

Section snippets

Mosquitos, mosquito saliva, and salivary gland extracts

Female Ae. vexans, Ae. triseriatus, Cx. tarsalis , and Culiseta (Cs.) inornata mosquitos were collected in local fields and identified by scientists in the Department of Entomology at the University of Manitoba. The Ae. aegypti colony was obtained from the same Department and maintained in our laboratory. The Cx. quinquefasciatus colony was imported from Dr. Robert J. Novak's laboratory at the University of Illinois (Champaign, Ill.) and maintained in our laboratory. Adult Ae. aegypti and Cx.

Salivary allergens of 10 mosquito species

As illustrated in Fig. 2 and Table II, immunoblot analysis revealed that there were 3 to 16 allergens in each of the 10 mosquito salivary extracts.

. Salivary allergens of 10 mosquito species. Proteins in saliva or salivary gland extracts of 10 species were separated by SDS-PAGE and then transferred to nitrocellulose membranes. Membranes were sequentially incubated with pooled serum from subjects allergic to mosquitos, monoclonal anti-human IgE, and enzyme-conjugated goat anti-mouse IgG as

Discussion

Mosquito salivary secretions are directly responsible for skin reactions to mosquito bites.27 Lack of commercially available, specific, high-quality mosquito saliva allergens has been a major rate-limiting step in the study of mosquito allergy to date. Collection of mosquito saliva or dissection of salivary glands is time-consuming and labor-intensive. By using these materials, it has been found that mosquito salivary gland–specific IgE is significantly increased in subjects allergic to

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    From athe Section of Allergy and Clinical Immunology, and bthe Department of Immunology, Faculty of Medicine, Department of Pediatrics and Child Health, University of Manitoba, Winnipeg.

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    Supported by the Children's Hospital Foundation, Winnipeg, Manitoba, Canada.

    Reprint requests: Zhikang Peng, MS746A-820 Sherbrook St., Winnipeg, Manitoba, Canada R3A 1R9.

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