Tick-host conflict: immunoglobulin E antibodies to tick proteins in patients with anaphylaxis to tick bite

Abstract

Tick-borne infectious diseases and allergies are a growing problem worldwide. Tick bite allergy has been associated with the direct effect of immunoglobulin E (IgE) response to tick salivary antigens, or secondary to the induction of allergy to red meat consumption through IgE antibodies against the carbohydrate α-Gal (Gal α 1-3Gal β 1-(3)4GlcNAc-R). However, despite the growing burden of this pathology, the proteins associated with anaphylaxis to tick bite have not been characterized. To address this question, a comparative proteomics approach was used to characterize tick proteins producing an IgE antibody response in a healthy individual with record of tick bites, which had not resulted in any allergic reactions, and two patients with anaphylactic reactions to Rhipicephalus bursa or Hyalomma marginatum tick bites. Both patients and the healthy individual were red meat tolerant. The results supported a patient-specific IgE antibody response to tick species responsible for the anaphylaxis to tick bite. Both patients and the healthy individual serologically recognized tick proteins with and without α-Gal modifications, with proteins differentially recognized by patients but not control sera. These proteins could be used as potential antigens for diagnostics, treatment and prevention of tick bite-induced allergies.

Keywords: Immune response; Immunity; Immunology and Microbiology Section; allergy; alpha-Gal; anaphylaxis; immunology; proteomics.

Figure 1. Case presentation for patient 1

A. The anaphylactic reaction was diagnosed in patient 1 after R. bursa tick bite that resulted in generalized itching, difficult breathing, nausea and somnolence that required medical attention. B. Female R. bursa attached to patient's skin and shown in more detail in the inset. C. Patient´s positive intradermal reaction to cetuximab (1:100 to 1:10 dilution; arrows). Intradermal reaction to cetuximab (1:1000 dilution) and to cetuximab prick were negative. Histamine prick was used as positive skin test control.

Figure 2. Immunological response to tick proteins

The IgE, IgM and IgG antibody levels were determined by ELISA in patients and control serum samples against A. α-Gal, B. R. bursa salivary gland proteins, C. H. marginatum salivary gland proteins, and D. R. microplus BME/CTVM23 tick cell proteins. Antibody levels were determined as OD at 450 nm and shown as average + SD of 4 technical replicates.

Figure 3. Patient-specific antibody response to tick species responsible for the reported anaphylactic reaction to tick bite

A. Correlation analysis between IgE, IgM and IgG antibody levels against R. bursa or H. marginatum tick proteins and α-Gal in patients 1 and 2 and healthy control individual. Antibody levels were determined as OD at 450 nm and shown as the average of 4 technical replicates. B. The IgE response to R. bursa and H. marginatum salivary gland and R. microplus BME/CTVM23 tick cell proteins was analyzed by 1-D Western blot using patient 1 and 2 sera. Abbreviation: MW, molecular weight protein marker.

Figure 4

Rhipicephalus tick proteins recognized by IgE in patient 1 and control sera and by anti-α-Gal IgE antibodies. The R. microplus BME/CTVM23 tick cell proteins were extracted and analyzed by 2-D Western blot using patients and control sera and anti-α-Gal antibodies. The protein spots of interest recognized by patients or control sera and by anti-α-Gal antibodies were manually excised from the stained gel and used for proteomics analysis. The same settings were used for all four panels in which proteins were resolved by isoelectrical focusing at pH 3-11 followed by 12% SDS gel electrophoresis in the second dimension with 140-15 kDa molecular weight range.

Figure 5

Hyalomma tick proteins recognized by IgE in patient 2 and control sera and by anti-α-Gal IgE antibodies. The H. marginatum salivary glands were dissected and proteins were extracted and analyzed by 2-D Western blot using patients control sera and anti-α-Gal antibodies. The protein spots of interest recognized by patient or control sera and by anti-α-Gal antibodies were manually excised from the stained gel and used for proteomics analysis. The same settings were used for all four panels in which proteins were resolved by isoelectrical focusing at pH 3-11 followed by 12% SDS gel electrophoresis in the second dimension with 140-15 kDa molecular weight range.

Figure 6. Gene ontology for

Rhipicephalus tick proteins recognized by IgE in patient 1 and control sera and by anti-α-Gal IgE antibodies. The R. microplus BME/CTVM23 tick cell proteins identified by proteomics analysis were functionally annotated for molecular function and biological process. A. Molecular function for unique proteins identified by patient and control sera. B. Biological process for unique proteins identified by patient and control sera. C. Molecular function for proteins recognized by anti-α-Gal antibodies. D. Biological process for proteins recognized by anti-α-Gal antibodies.

Figure 7. Gene ontology for

Hyalomma tick proteins recognized by IgE in patient 2 and control sera and by anti-α-Gal IgE antibodies. The H. marginatum salivary gland proteins identified by proteomics analysis were functionally annotated for molecular function and biological process. A. Molecular function for unique proteins identified by patient and control sera. B. Biological process for unique proteins identified by patient and control sera. C. Molecular function for proteins recognized by anti-α-Gal antibodies. D. Biological process for proteins recognized by anti-α-Gal antibodies.