Identification of major and minor chicken allergens in dogs.

BACKGROUND: Allergens targeted by serum-specific immunoglobulin E (sIgE) in dogs clinically allergic to chicken have not been reported. OBJECTIVES: To characterise the allergens targeted by sIgE in dogs sensitised and allergic to chicken. ANIMALS: Sera from three dogs not sensitised to chicken, from 10 chicken sensitised dogs and from 12 chicken allergic dogs. METHODS AND MATERIALS: Enzyme-linked immunosorbent assay (ELISA) and immunoblotting with a commercial chicken extract were utilized. The bands identified on immunoblotting were sequenced by mass spectrometry for allergen characterization. RESULTS: Using ELISA, we detected chicken-sIgE above the positive threshold in zero of three (0%) nonsensitised dogs, five of five (100%) chicken-sensitised dogs (a selection criterion), and in seven of 12 (58%) chicken-allergic dogs. Immunoblotting performed with the same extract revealed IgE-bound protein bands in 100% of all chicken-sensitised and -allergic dogs, respectively. To identify the allergens, we excised the corresponding bands on the electrophoretic gel, and submitted them for sequencing by mass spectrometry. We conclusively identified seven major allergens (serum albumin, pyruvate kinase M, enolase 3, creatine kinase M, lactate dehydrogenase A, glyceraldehyde-3-phosphate dehydrogenase and triose-phosphate isomerase) and one minor allergen (troponin C), which are relevant to dogs. CONCLUSIONS AND CLINICAL RELEVANCE: We identified herein seven major chicken allergens for dogs, several of which are known to be cross-reactive allergens for humans. Based on their degree of sequence identity, these allergens exhibit the theoretical potential to be cross-reactive between poultry and mammalian meats; six of these allergens already are known to be cross-reactive between chicken and fish species. Future studies should address the clinical relevance and cross-reactivity potential of these chicken allergens in dogs.

Serum IgE cross-reactivity between fish and chicken meats in dogs.

BACKGROUND: In humans, a cross-reactive clinical allergy has been reported between three chicken and fish meat proteins: beta-enolase, aldolase A and parvalbumin. OBJECTIVE: To evaluate if IgE cross-reactivity between chicken and fish also existed in the dog. ANIMALS: Sera from dogs with suspected allergic skin disease and with IgE against chicken and fish. METHODS AND MATERIALS: Sera were analysed by ELISA and immunoblotting with chicken, white fish (haddock and cod) and salmon extracts. Reciprocal inhibition ELISAs and inhibition immunoblots were then performed. Protein sequencing of bands identified on multiple extracts was determined by mass spectrometry. RESULTS: Out of 53 archived canine sera tested by ELISA against chicken, white fish or salmon, 15 (28%), 12 (23%) and 26 (49%), respectively, had elevated IgE against one, two or all three of these extracts. Seven of the triple-reactive sera were subjected to reciprocal inhibition ELISAs. A >50% inhibition was found between chicken-fish, chicken-salmon and fish-salmon in seven, four and five of seven dogs, respectively. Immunoblotting identified multiple IgE-binding proteins of identical molecular weights in the three extracts; these were partially to fully cross-reactive by inhibition immunoblotting. Mass spectrometry identified nine cross-reactive proteins as: pyruvate kinase, creatine kinase, alpha-actin, glyceraldehyde-3-phosphate dehydrogenase, beta-enolase, aldolase, malate dehydrogenase, lactate dehydrogenase and triose-phosphate isomerase 1. All of these have been reported previously as fish, shellfish and/or chicken allergens for humans. CONCLUSIONS AND CLINICAL IMPORTANCE: Whether any of these newly identified IgE cross-reactive chicken-fish allergens is the cause of clinical allergy needs to be determined in dogs reacting to at least two of these common food sources.

Cornstarch is less allergenic than corn flour in dogs and cats previously sensitized to corn.

BACKGROUND: Corn appears to be an uncommon food source of allergens in dogs and cats. There is limited information on the nature of the corn allergens in dogs and cats and their presence in the various foodstuffs used in commercial pet foods. The aim of this study was to determine if serum IgE from corn-sensitized dogs and cats recognized proteins in corn flour and cornstarch, which are common sources of carbohydrates in pet foods. RESULTS: We selected archived sera from allergy-suspected dogs (40) and cats (40) with either undetectable, low, medium or high serum levels of corn-specific IgE. These sera were tested then by ELISA on plates coated with extracts made from corn kernels, corn flour, cornstarch and the starch used in the commercially-available extensively-hydrolyzed pet food Anallergenic (Royal Canin). Immunoblotting was then performed on the same extracts with some of the sera from moderate-to-high corn-sensitized dogs and cats. Using ELISA, it is mostly the dogs and cats with moderate and high corn-specific IgE levels that also had IgE identifying allergens in the flour (dogs: 20/30 sera, 67% - cats: 20/29, 69%). In contrast, none of the tested sera had measurable IgE against proteins isolated from the cornstarch. Immunoblotting confirmed the existence of numerous major corn allergens in the corn kernel extract, fewer in that of the corn flour, while such allergens were not detectable using this technique in the two cornstarch extracts. CONCLUSIONS: In this study, ELISA and immunoblotting results suggest that IgE from corn-sensitized dogs are less likely to recognize allergens in cornstarch than in kernel and flour extracts. As corn is not a common allergen source in dogs and cats, and as its starch seems to be less allergenic than its flour, pet foods containing cornstarch as a carbohydrate source are preferable for dogs and cats suspected of suffering from corn allergy.

Extensive protein hydrolyzation is indispensable to prevent IgE-mediated poultry allergen recognition in dogs and cats.

BACKGROUND: The central premise for the commercialization of diets with hydrolyzed ingredients is that the small-sized digested peptides would be unable to crosslink allergen-specific IgE at the surface of tissue mast cells and induce their degranulation. Evidence for the validity of this concept to diagnose food allergies in dogs and cats is limited, however. Our objectives were to study the recognition of standard and variably hydrolyzed poultry extracts by sera from dogs and cats with elevated chicken-specific serum IgE. RESULTS: Forty sera from dogs and 40 from cats with undetectable, low, medium or high serum levels of chicken-specific IgE were tested by ELISA on plates coated with the positive controls chicken, duck and turkey meat extracts and the negative controls beef meat (dogs) or wheat (cats). Plates were also coated with a non-hydrolyzed chicken meal, and mildly- or extensively-hydrolyzed poultry feather extracts. The frequencies of dogs with positive IgE against the various extracts were: chicken meat: 100%, duck and turkey meats: 97%, beef meat: 3%, non-hydrolyzed chicken meal: 73%, mildly-hydrolyzed poultry feathers: 37% and extensively-hydrolyzed poultry feathers: 0%. For cats, these respective percentages were (with wheat replacing beef as a negative control): 100, 84, 97, 7, 7, 0 and 0%. To detect any allergenic cross-reactivity between poultry meat-based and feather hydrolysate-derived extracts, an IgE ELISA inhibition was also done. Ten canine sera with the highest level of anti-poultry IgE in the previous experiment were incubated overnight with a previously optimized 50 µg amount of each of the extracts used above. We performed ELISA on plates coated with chicken, duck or turkey meats with or without inhibitors. The median inhibition percentages after incubation with the non-hydrolyzed chicken meal were ~22%, with the mildly-hydrolyzed poultry feathers: 14-22%, and those with the extensively-hydrolyzed poultry feathers: 5 to 10%; the last inhibition level was similar to that of the beef meat negative control. CONCLUSIONS: Altogether, these results suggest that an extensive-but not partial-hydrolyzation of the poultry feather extract is necessary to prevent the recognition of allergenic epitopes by poultry-specific IgE.

Co-sensitization and cross-reactivity between related and unrelated food allergens in dogs - a serological study.

BACKGROUND: Knowledge of cross-reactivity between foods is useful so that potentially cross-reactive allergens can be avoided in diet trials. HYPOTHESIS/OBJECTIVES: To evaluate allergenic cross-reactivity in related foods. ANIMALS: Sera from 469 dogs with suspected adverse food reactions. METHODS: An IgE-based serological assay using 19 food allergens was performed in 469 dogs. Pairwise comparisons were used to calculate the odds ratios (ORs) for each food pair, with significance at P < 0.0002 by Holm-Bonferroni correction, both in all 469 dogs and in the 261 of 469 dogs with at least one positive reaction. One-way ANOVA with Tukey's post hoc tests (significance at P < 0.05) were used to test for differences between mean logE ORs in different food groups. Inhibition enzyme-linked immunosorbent assays (ELISAs) were performed to assess allergenic cross-reactivity between beef, lamb and cow's milk. RESULTS: Significant associations were observed between both related and unrelated food pairs. Associations were, however, more frequent and stronger among related than unrelated foods. In all 469 dogs, 38 of 43 related food pairs were significantly associated [mean (SD) logE OR 3.4 (0.9)] compared with 79 of 128 unrelated pairs [2.7 (1.0)], P < 0.0002. In positive dogs, 32 of 43 related pairs were significantly associated [2.7 [1.0)] compared with 49 of 128 unrelated pairs [1.8 (1.0)], P < 0.0002. Inhibition ELISAs confirmed the presence of cross-reactive IgE-binding epitopes in beef, lamb and cow's milk. CONCLUSIONS AND CLINICAL IMPORTANCE: The results suggest that related and potentially cross-reactive foods should be avoided in elimination diets.

Serum anti-Staphylococcus pseudintermedius IgE and IgG antibodies in dogs with atopic dermatitis and nonatopic dogs.

BACKGROUND: Dogs and humans with atopic dermatitis (AD) are predisposed to colonization and recurrent infection with Staphylococcus spp. Studies in humans suggest that staphylococcus-specific immunoglobulin E (IgE) plays a key role in disease pathogenesis. Few such studies have been undertaken in dogs. HYPOTHESIS/OBJECTIVES: The aim of this study was to compare levels of staphylococcus-specific IgE and immunoglobulin G (IgG) in dogs with AD, nonatopic dogs with staphylococcal pyoderma, and nonatopic and noninfected control dogs. ANIMALS: Sera were collected from 108 dogs with AD, 39 nonatopic dogs with staphylococcal pyoderma secondary to different underlying conditions, 67 age-matched nonatopic control dogs, and nine control dogs reared in minimal disease conditions. METHODS: Serum Staphylococcus pseudintermedius-specific IgE and IgG antibodies were measured by enzyme-linked immunosorbent assay. RESULTS: Dogs with AD had significantly higher levels of anti-staphylococcal IgE than nonatopic dogs with staphylococcal pyoderma and the two groups of control dogs. Levels of anti-staphylococcal IgG were significantly higher in atopic dogs and nonatopic dogs with pyoderma compared with nonatopic control dogs and control dogs reared in minimal disease conditions, but there was no significant difference in levels of anti-staphylococcal IgG between dogs with AD and nonatopic dogs with pyoderma. CONCLUSIONS AND CLINICAL IMPORTANCE: A significantly increased IgE response to S. pseudintermedius antigens in atopic dogs suggests an immunopathogenic role for anti-staphylococcal IgE. The finding of elevated IgE and IgG in atopic dogs is also important as a prelude to studies on antigenic specificity and possible correlations with disease phenotype.

Patch testing and allergen-specific serum IgE and IgG antibodies in the diagnosis of canine adverse food reactions.

Adverse food reaction (AFR) is a common differential diagnosis for pruritic dogs. The only way to diagnose AFR is an elimination diet of 6-8 weeks with a protein and a carbohydrate source not previously fed. In humans, patch testing has been shown to be a useful tool to diagnose food allergies. In veterinary medicine, serum food allergen-specific antibody testing is widely offered to identify suitable ingredients for such diets. The aim of this study was to determine sensitivity, specificity, negative and positive predictability of patch testing with and serum antibody testing for a variety of common food stuffs. Twenty-five allergic dogs underwent an elimination diet and individual rechallenge with selected food stuffs, food patch testing and serum testing for food-antigen specific IgE and IgG. Eleven clinically normal control dogs only were subjected to patch and serum testing. The sensitivity and specificity of the patch test were 96.7 and 89.0% respectively, negative and positive predictability were 99.3 and 63.0%. For IgE and IgG the sensitivity was 6.7 and 26.7%, specificity were 91.4 and 88.3%, the negative predictive values 80.7 and 83.7% and the positive predictive values were 15.4 and 34.8%. Based on these results, a positive reaction of a dog on these tests is not very helpful, but a negative result indicates that this antigen is tolerated well. We conclude that patch testing (and to a lesser degree serum testing) can be helpful in choosing ingredients for an elimination diet in a dog with suspected AFR.

Food allergen-specific serum IgG and IgE before and after elimination diets in allergic dogs.

Serum food allergen-specific antibody testing is widely offered to identify suitable ingredients for diets to diagnose adverse food reaction (AFR) in dogs with allergic skin disease. Antibody concentrations in blood samples obtained during an unsuccessful diet to help in the choice of diet changes may be influenced by the previous diet. The objective of this paper was to measure food antigen-specific IgE and IgG for the most commonly used 16 food antigens before and after an elimination diet. Levels of food-specific serum IgE and IgG antibodies were measured by enzyme-linked immunosorbent assay (ELISA). Dogs had detectable IgE antibodies to beef, pork, lamb and cows' milk; and detectable IgG antibodies to beef, pork, lamb, cows' milk, chicken and turkey. Of 19 dogs with complete data sets, 14 dogs showed clear improvement during diet and in 7 dogs AFR could be diagnosed by deterioration on rechallenge and subsequent improvement on refeeding the diet. Serum was obtained before and 6-8 weeks after beginning such a diet. There was no significant difference in pre- and post-diet levels for any of the individual allergens nor for the total IgE and IgG concentrations of all antigens (P=0.55 and P=0.53 respectively). In these 19 dogs in which an elimination diet was used for the diagnosis of food allergy and in which 14 were probably food allergic and 7 were proven food allergic there were no significant differences in food-specific antibodies before and after an elimination diet of 6-8 weeks.

Levels of house dust mite-specific serum immunoglobulin E (IgE) in different cat populations using a monoclonal based anti-IgE enzyme-linked immunosorbent assay.

Levels of serum immunoglobulin E (IgE) specific for the house dust mites (HDMs) Dermatophagoides farinae (DF) and Dermatophagoides pteronyssinus (DP) in 58 cats with clinical signs suggestive of atopic dermatitis (allergic dermatitis cats), 52 cats with no history of allergic or immunological disease (nonallergic cats) and 26 specific pathogen-free (SPF) cats were measured using a monoclonal anti-IgE enzyme-linked immunosorbent assay. Reactivity to both native and reduced HDM allergens was compared. SPF cats had significantly lower levels of HDM-specific serum IgE than cats with allergic dermatitis and nonallergic cats. The difference in levels of HDM-specific IgE in the serum of cats with allergic dermatitis and nonallergic cats was significant for native DF allergen, but not for native DP allergen or reduced HDM allergens. The results suggest that DF in its native form may be a significant allergen in cats with allergic dermatitis. The clinical relevance of these reactions, however, remains to be proven.