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Keywords:

  • allergic asthma;
  • bird-egg syndrome;
  • chicken albumin;
  • egg allergy;
  • feathers

Abstract

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References

Background: Chicken serum albumin (α-livetin) has been implicated as the causative allergen of the bird-egg syndrome. However, the clinical relevance of sensitization to this allergen has not been confirmed by specific challenge tests and environmental sampling. We investigated whether chicken albumin can be detected in air samples collected in a home with birds, and whether sensitization to this protein may cause respiratory and food allergy symptoms. The heat resistance of chicken albumin and the possible cross-reactivity with conalbumin were also investigated.

Methods: We studied eight patients with food allergy to egg yolk who also suffered from respiratory symptoms (rhinitis and/or asthma) caused by exposure to birds. Sensitization to egg yolk and bird antigens was investigated by skin and serologic tests. Hypersensitivity to chicken albumin was confirmed by specific bronchial, conjunctival, and oral provocation tests.

Results: All patients had positive skin tests and serum IgE against egg yolk, chicken serum, chicken meat, bird feathers, and chicken albumin. The presence of airborne chicken albumin in the domestic environment was confirmed. Specific bronchial challenge to chicken albumin elicited early asthmatic responses in six patients with asthma. An oral challenge with chicken albumin provoked digestive and systemic allergic symptoms in the two patients challenged. IgE reactivity to chicken albumin was reduced by 88% after heating at 90°C for 30 min. ELISA inhibition demonstrated only partial cross-reactivity between chicken albumin and conalbumin.

Conclusions: Chicken albumin (Gal d 5) is a partially heat-labile allergen that may cause both respiratory and food-allergy symptoms in patients with the bird-egg syndrome.

In recent years, a relationship between respiratory type I hypersensitivity to bird antigens and food allergy to egg yolk has been described (1–5). Egg-yolk α-livetin, also known as chicken serum albumin (CSA), has been reported to be the cross-reacting allergen responsible for this association (5–7). It is thought that sensitization to airborne bird allergens precedes allergy to egg antigens. This phenomenon may be considered a paradigm of class 2 food allergy, which is mainly seen in adults and develops as a consequence of allergic sensitization to inhalant allergens (8). To the best of our knowledge, the ability of CSA to elicit asthma symptoms has only been documented as a case report by our group (7). No other studies have demonstrated the clinical relevance of sensitization to CSA by means of inhalation or oral challenge tests.

The pathogenic mechanism of bird-egg syndrome is different from pigeon fancier's lung, a form of hypersensitivity pneumonitis caused mainly by immunologic responses to IgA and mucin contained in feather bloom and pigeon droppings (9). Although hypersensitivity pneumonitis is the most commonly reported presentation following sensitization to bird antigens (9, 10), asthmatic responses have also been described (11, 12). Moreover, inhalation of several egg proteins has been reported as a cause of allergic asthma (7, 13–18). Little is known, however, about the properties of the antigenic components in feathers that may cause IgE-mediated sensitization and asthma.

In this paper, we describe eight patients who experienced rhinoconjunctivitis, with or without asthma symptoms, upon exposure to birds. These patients also suffered from allergy symptoms after the ingestion of eggs. Specific bronchial and oral challenge provocation tests to CSA were carried out to confirm the causative role of this protein in the clinical manifestations of the bird-egg syndrome. We also investigated whether CSA can be detected as an aeroallergen in the home environment of patients with birds. The resistance of CSA to heating was assessed, since patients suffering from the bird-egg syndrome may tolerate the ingestion of well-cooked eggs and poultry.

Material and methods

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References

Patients

Eight patients (five men/three women) who suffered from respiratory symptoms (all of them from rhinoconjunctivitis and six from asthma) upon exposure to bird feathers, as well as allergy symptoms after ingestion of egg yolk, were studied in 1998–9 in our allergy clinic. These patients ranged in age between 21 and 41 years (mean, 30.75 years). The clinical and demographic data of the patients are shown in Table 1. Patients 3 and 4 were brothers. Specific questions were asked of each subject about the presence of birds at home and eventual symptoms after egg ingestion or contact with birds. These patients had frequent or sporadic contact with some type of birds (budgerigars, parrots, canaries, pigeons, and hens), mostly at home but, occasionally, away from home. At the time of the study, four patients kept birds as household pets (budgerigars or canaries). In these subjects, cleaning the bird's cage provoked rhinoconjunctivitis and asthma attacks within a few minutes. Two patients had removed their birds several years before. Interestingly, two of the patients reported respiratory symptoms after contact with a flock of feral city pigeons. One patient reported severe asthma attacks when he visited a pet shop; another experienced asthma symptoms while working as a construction worker at a poultry farm. The onset of respiratory symptoms with birds preceded egg allergy in four patients, and in the remaining four patients there was a simultaneous onset. None of them showed the clinical features of hypersensitivity pneumonitis.

Table 1.  Demographic and clinical data of study population
PatientSex Age (years)Symptoms afterPeriod (years) since onset of symptomsBird contact at home or away*Previous contact with birds
Bird contactEgg ingestionPoulty ingestionBird contactEgg ingestion
  1. conjunct.: conjunctivitis; OAS: oral allergy syndrome; AE: angioedema, CU: contact urticaria.

1M/38Rhinitis, conjunct., asthmaOAS, AE, asthma, anaphylaxisTolerate cooked4112 years with 4 canaries, *poultry farms
2F/41Rhinitis, conjunct., asthmaOAS, abdominal painTolerate cooked10.52 years with 1 budgerigar; *wild pigeons
3M/25Rhinitis, conjunct.OAS, AECU raw; tolerate cooked>20>20Since birth, *hens, pigeons
4M/27Rhinitis, conjunct.OAS, AECU raw; tolerate cooked>20>20Since birth, *hens, pigeons
5F/31Rhinitis, conjunct., asthmaOAS, AE, conjunct.CU raw; tolerate cooked1211*Wild pigeons2 canaries, 12 years before for 3 years
6F/29Rhinitis, conjunct., asthmaOAS, AE, asthmaCU raw; tolerate cooked10101 parrot, 10 years before for 5 years
7M/34Rhinitis, conjunct., asthmaOASTolerate cooked223 years with 2 budgerigarsSince birth, budgerigars, canaries
8M/21Rhinitis, conjunct., asthmaOAS, AETolerate cooked435 years with 2 canaries, *pet shopsSince birth, canaries

All patients experienced itching and burning of the mouth immediately after eating fried egg yolk (usually undercooked), and sometimes with homemade mayonnaise. This was followed by swelling of the lips and oral mucosa, and sometimes facial angioedema. Patients 1 and 6 also suffered shortness of breath and chest tightness after egg ingestion. Most patients tolerated well-cooked eggs, such as boiled eggs and omelets. Patients 3, 4, 5, and 6 also experienced pruritic wheals on their hands and faces shortly after handling raw chicken meat or fresh hen's eggs. Ingestion or manipulation of well-cooked chicken produced no symptoms in any patient.

Written informed consent was obtained from all patients to be included in the study.

Allergen extracts

Livetins were extracted from fresh hen's egg yolk as previously described (15), lyophilized, and reconstituted at a final concentration of 2 mg/ml in phosphate-buffered saline (PBS). The protein concentration of this extract, as estimated by the Lowry-Biuret method (Sigma Chemical Co., St Louis, MO, USA), was 48% w/w. Ovomucoid (T-2011), ovalbumin (A-5378), conalbumin (C-0755), lysozyme (L-6876), and chicken serum albumin (fraction V powder, A-3014) were purchased from Sigma Chemical Co. and prepared at a concentration of 10 mg/ml in PBS. CSA was purchased at the highest available purification level (99% purity by agarose electrophoresis and globulin-free). All these extracts were passed through a 22-μm filter (Millipore Corp., Bedford, MA, USA) for sterilization.

Skin tests

Skin prick tests were performed with a battery of common inhalant allergens, including Dermatophagoides pteronyssinus and D. farinae; grass, weed, and tree pollens; molds; and dog and cat dander (C.B.F. LETI SA, Madrid, Spain). Commercially available extracts of whole egg (Bencard, Worthing, UK), egg white, egg yolk, feather mix (chicken and duck), chicken meat (ALK-Abelló, Madrid, Spain), and chicken serum (C.B.F. LETI SA), as well as purified ovomucoid, ovalbumin, conalbumin, lysozyme, and CSA (Sigma Chemical Co.), were tested. In addition, skin end-point titration with CSA was done by testing twofold concentrations of the extract by the prick method. Histamine phosphate at 10 mg/ml and normal saline were used as positive and negative controls, respectively. The response was read 15 min after puncture, and the results were expressed as the mean wheal diameter (mm). A wheal diameter 3 mm or greater with erythema, compared with the saline control, was defined as a positive reaction. Skin tests were examined again 6 h later to assess a possible late skin reaction (erythema and swelling). Skin prick tests with the aforementioned egg and avian extracts were also performed in 10 atopic and 10 nonatopic control subjects.

Bronchial provocation tests

The methacholine inhalation test was performed according to Cockcroft et al. (19), with some modifications. The aerosolized particles were generated by a continuous pressurized nebulizer model De Vilbiss 646 (De Vilbiss Co., Somerset, PA, USA) with an output of 0.28 ml/min. The result of this test was expressed as the provocative concentration of methacholine causing a 20% fall in forced expiratory volume in 1 s (PC20), and it was determined by interpolation of the last two concentrations. This test was performed in five patients with asthma, but not in patient 7, who had baseline airways obstruction and a positive bronchodilator test to salbutamol inhalation.

Specific bronchial challenge to CSA was carried out in the six patients with asthma by the method previously described (7). The patient inhaled the aerosolized allergen by the nebulizer method mentioned above in progressive concentrations at tidal breathing for 2 min. A control challenge with PBS was carried out before antigen provocation. Increasing concentrations of CSA extract were given by inhalation, starting with a concentration that induced a 2–3-mm wheal on skin prick testing. The dose was increased in twofold increments at 10-min intervals, and FEV1 was measured at 5 and 10 min after inhalation of each concentration. The bronchial challenge test was discontinued when there was a fall in FEV1 of 20% or greater from the lowest postsaline value, or when the highest concentration had been given. At the end of the inhalation challenge test, spirometry was performed at 20, 30, 40, and 60 min after challenge, and again the following day. Peak expiratory flow was measured before and every hour after bronchial challenge until bedtime, and again the day after. A fall in FEV1 of 20% or more from the lowest postsaline value within 60 min of challenge was considered a positive immediate reaction, while a fall in peak expiratory flow greater than 25% at 2–24 h after challenge was considered a positive late reaction if no change was observed during the control day. PC20 allergen was calculated as described above.

Conjunctival challenge test

The conjunctival challenge test was performed as described by Möller et al. (20), with twofold increasing concentrations of the CSA extract. This test was carried out in patients 3 and 4, who had only rhinoconjunctivitis symptoms. Two atopic patients were also tested as control subjects.

Oral challenge tests

A double-blind, placebo-controlled, oral challenge test with CSA was carried out as described elsewhere (21) in patients 1 and 5, who gave informed consent. CSA was reconstituted in normal saline at a concentration of 20 mg/ml, and several twofold dilutions were prepared. One milliliter of either placebo (normal saline) or freshly prepared solutions of CSA containing 0.16, 0.31, 0.62, 1.25, 2.5, 5, and 10 mg/ml was administered orally. The dose was doubled at 30-min intervals until the patient experienced allergic symptoms, or a maximum amount of 20 mg of CSA was reached. Spirometry was performed before each oral challenge and at 10-min intervals for the first 30 min after the challenge. Patients were observed for the development of respiratory, skin, or gastrointestinal symptoms for 2 h after the last dose was administered.

Total and specific IgE determination

Total serum IgE was measured by the Pharmacia CAP System IgE FEIA (Pharmacia, Uppsala, Sweden) according to the manufacturer's instructions. Specific IgE against egg yolk, egg white, ovalbumin, ovomucoid, conalbumin, lysozyme, feather mix (budgerigar, canary, parrot), chicken feathers, and chicken meat was determined by Pharmacia CAP System RAST FEIA. A result higher than 0.35 kU/l was regarded as positive.

The serum levels of specific IgE against CSA and livetins were determined in duplicate by direct ELISA, as described elsewhere (22), with plastic microtiter plates (Immulon 4, Chantilly, VA, USA) coated at a concentration of 1 µg of protein/well and serum dilutions starting from 1:4. A result was considered positive when a serum bound four times more specific IgE than the mean titer of four control subjects allergic to egg white, but not to egg yolk.

The resistance of the egg proteins to heat treatment was assessed by direct ELISA using unheated and heated (90°C for 30 min) livetin, CSA, and conalbumin extracts. These assays were performed with a serum pool of equal-volume aliquots of sera from patients 1 to 7.

ELISA-inhibition assays

Competitive ELISA-inhibition assays were conducted to investigate the possible existence of allergenic cross-reactivity between CSA and conalbumin. The heat resistance of CSA and livetin extracts was also assessed by ELISA inhibition. The serum pool from patients 1 to 7 was used. Briefly, 50 µl of progressive twofold dilutions of the inhibitor allergens or 50 µl of PBS, as a negative control, was preincubated with 50 µl of the serum pool for 2 h, and then the assay was continued by standard ELISA-inhibition techniques. Unheated CSA was used as the immobilized allergen; livetins and CSA (both unheated and heated) and unheated conalbumin were used as the aqueous competitive allergens. Results were expressed as the percent inhibition of IgE ELISA obtained with the inhibitor allergen as compared with a control assay with PBS inhibitor.

SDS–PAGE and immunoblotting

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS–PAGE) was carried out by Laemmli's method in reducing conditions (23). Polyacrylamide concentrations of 12% T and 2.67% C were used for the separating gels. An amount of 10 µg of protein was applied per lane. Bio-Rad standards were used as molecular weight (MW) markers (Bio-Rad Lab., Hercules, CA, USA). The protein bands were visualized with Coomassie brilliant blue R-250. The separated proteins were electrotransferred to Immobilon-P membranes (Millipore, Bedford, MA, USA) by the method of Towbin (24). Unreacted membrane sites were blocked with PBS containing 3% human serum albumin and 0.05% Tween-20. The membrane was then incubated with the patient's serum diluted 1:50 in 10 mM PBS containing 0.1% Tween-20 (PBS-T). After washing with PBS-T, the membranes were incubated with monoclonal antihuman IgE conjugated with peroxidase at a concentration of 1 µg/ml (Ingenasa SA, Madrid, Spain). The protein bands were developed with 3-3′,5-5′ tetramethylbencidine and oxygen peroxide as substrate.

Immunoblotting inhibition

The specificity of IgE binding to CSA was investigated by immunoblot inhibition. Assays were performed with a serum pool from patients 1 to 7 preincubated for 12 h with equal volumes of unheated CSA, livetin, or chicken feather extracts, followed by immunodetection as described above.

Air sampling and extraction of air filters

Air samples were collected from a dwelling at a single location in a 19 m3 room where 11 free-flying budgerigars were kept. No other pets were kept in this household. A volumetric air sampler (Quan-Tec-Air, Inc, Rochester, MN, USA) was used to collect air samples (25). The air sampler was operated 12 h/day at a flow rate of 2.5 l/s on eight different days. Airborne particles were collected onto polytetrafluoroethylene filters (Quan-Tec-Air, Inc). The allergens were directly extracted from the air samples by removing the filter from the woven backing and eluting it overnight in 1 ml of PBS containing 1% Tween-20. The extracts were centrifuged and the supernatants stored at −20°C until they were analyzed.

Precipitating antibodies

Precipitating antibodies against feathers, serum, and droppings from budgerigars and pigeons (Bial-Aristegui, Bilbao, Spain) were determined by double diffusion according to Ouchterlony (26).

Results

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References

Skin tests

Skin prick tests results are shown in Table 2. All patients showed positive skin responses to egg yolk, CSA, feathers, chicken serum, and chicken meat. Three patients showed positive skin responses to egg white, and six patients to conalbumin. Seven out of the eight patients had negative skin responses to the main egg white allergens: ovalbumin, ovomucoid, and lysozyme. All patients showed late skin reactions (erythema and swelling) to CSA and chicken serum 6 h after skin testing. Skin prick tests to these egg and avian-derived extracts in the 10 atopic and 10 nonatopic control subjects were negative, the differences being statistically (chi-square) significant as compared with the patient group (P<0.00001).

Table 2.  Skin prick test results (mean wheal in mm)
Patients12345678
  1. HDM: house-dust mites; Neg.: negative.

Egg whole 5% w/v86577885
Egg yolk 5% w/v1065851075
Egg white 5% w/v50004060
Ovomucoid 10 mg/ml00000080
Ovalbumin 10 mg/ml00000000
Conalbumin 10 mg/ml604665100
Lysozyme 10 mg/ml00000060
Chicken albumin 10 mg/ml19615201110128
Chicken serum 50% v/v156141281085
Chicken meat 5%66475784
Feather mix 5%5771041375
Positive skin tests to common aeroallergensHDM, dogPollenPollen, HDM, catPollen, HDMNeg.Pollen, cat, dogPollen, dog, AlternariaPollen

Bronchial provocation tests

Inhalation challenge with methacholine revealed bronchial hyperresponsiveness in three out of the five patients tested. The bronchial provocation test to CSA extract elicited early asthmatic responses in the six patients with asthma symptoms. No late asthmatic responses were observed. The bronchial provocation test to CSA was negative in two control asthmatic subjects. The results of bronchial provocation tests are shown in Table 3.

Table 3.  Results of bronchial provocation tests (BPT) to methacholine and CSA
PatientsMethacholine inhalation test PC20 (mg/ml)BPT to CSA Maximum fall in FEV1 from baseline (%)BPT to CSA PC20 allergen (mg/ml)
  1. ND: not done.

1>16250.008
20.20260.034
5>16210.009
60.90250.012
7ND280.003
80.13230.015
2 control subjectsNeg.

Conjunctival challenge test

Conjunctival challenge with CSA elicited an intense reaction (ocular itching and redness) in patients 3 and 4 at concentrations of 0.078 and 0.039 mg/ml, respectively. This test was negative in two control atopic subjects.

Oral challenge tests

Ten minutes after oral administration of 15 mg of CSA, patient 1 experienced severe ocular injection, chemosis, and ocular itching, as well as angioedema of the oral mucosa, tongue, and eyelids. Patient 5 suffered intense itching in the oropharynx and ears 5 min after 10 mg of CSA was administered, followed by abdominal pain and coughing 10 min after the challenge. Allergic symptoms subsided after treatment with antihistamines and systemic corticosteroids in both patients.

Total and specific IgE determination

The results of total serum IgE and specific IgE determinations to egg and avian allergens are shown in Table 4. All patients showed a positive IgE determination to egg yolk and feathers, as well as to CSA and livetins. Negative results to these allergens were obtained in four control subjects allergic to egg white. After heating, IgE reactivity to CSA, livetins, and conalbumin extracts was reduced by 88%, 77.1%, and 26%, respectively.

Table 4.  Results of total and specific IgE to egg and avian allergens (kU/l), livetins and CSA (OD)
Patients12345678
  1. *CSA and livetin ELISA results expressed in OD at 490 nm. ELISA results to CSA in four control subjects allergic to egg white: 0.015±0.008 OD.

Egg yolk11.745.11.215.371.8023.482.40.61
Egg white5.2116.10.852.711.5911.420.9<0.35
Ovomucoid<0.35<0.35<0.35<0.35<0.35<0.351.25<0.35
Ovalbumin<0.35<0.35<0.35<0.35<0.35<0.35<0.35<0.35
Conalbumin<0.35<0.35<0.352.891.4218.223.3<0.35
Lysozyme<0.35<0.350.470.67<0.35<0.35<0.35<0.35
Chicken meat3.1637.50.994.181.3519.648.8<0.35
Feathers mix2.712.110.351.390.567.427.181.04
Chicken feathers3.66561.094.331.4621.429.71.05
Livetins*1.173.420.471.990.953.213.640.40
CSA*2.733.270.382.350.613.263.390.38
Total serum IgE14837765.460224353280188.2

ELISA-inhibition assays

The results of ELISA-inhibition experiments, shown collectively in Fig. 1, demonstrate that IgE binding to CSA could be inhibited in a dose-dependent manner up to 98% with unheated CSA and up to 88% with unheated livetin extract as liquid-phase allergens. The maximum inhibitions achieved with the heated CSA and livetin extracts were 87% and 45%, respectively. The maximum inhibition of IgE reactivity to CSA obtained with conalbumin was 53% at the highest concentration tested. The amount of protein necessary to achieve 50% inhibition in these assays was 0.001 µg for unheated CSA, 0.02 µg for unheated livetin, 0.025 µg for heated CSA, 36.7 µg for heated livetin, and 8.4 µg for conalbumin. These results indicate that heating significantly reduced the allergenicity of CSA and livetin extracts.

image

Figure 1. ELISA-inhibition assays using unheated CSA in solid phase and following inhibitors: unheated chicken serum albumin (CSA, 4°C), unheated livetins (LIVET, 4°C), heated chicken serum albumin (CSA, 90° C), heated livetins (LIVET, 90°C), and unheated conalbumin (CNA, 4°C).

Download figure to PowerPoint

Immunoblotting

Immunoblots with individual serum from patients 1 to 7 showed an IgE-binding band at 65–70 kDa in the extract of CSA (Fig. 2, lanes 1–7). IgE binding was weak in patients 3 and 5, in agreement with ELISA results. Serum from control subjects did not recognize this protein. IgE binding to CSA could be inhibited by preincubation of the serum pool (patients 1 to 7) with CSA 1 mg/ml (lane 8), livetin extract 1 mg/ml (lane 9), chicken feathers 1 mg/ml (lane 10), and air samples extract (lane 11), but not with normal saline (lane 12).

image

Figure 2. Lanes 1–7) immunoblots of CSA with individual serum from patients 1 to 7. Lanes 8–11) inhibition of CSA immunoblots with following inhibitor allergens (1 mg/ml): CSA (lane 8); livetin extract (lane 9); chicken feathers (lane 10); pool of air-sample extracts (lane 11). Lane 12) inhibited with normal saline. Lane 13) negative control serum.

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Immunoblot of air samples

The presence of CSA in most of the air samples collected from a house with 11 free-flying budgerigars was confirmed by immunoblotting (Fig. 3), an IgE-binding band at 66 kDa being detected. This band could be inhibited by preincubation of the pooled serum with CSA 1 mg/ml (lane 9) and chicken feathers 1 mg/ml (lane 10), as shown in Fig. 3.

image

Figure 3. Lanes 1–8) IgE binding to 66-kDa band in eight different air-sample extracts. Lanes 9 and 10) pool of air samples inhibited with CSA (lane 9) and chicken-feather extract (lane 10). Lanes 11 and 12) IgE binding to CSA (lane 11) and livetin extract (lane 12).

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Precipitating antibodies

No precipitating antibodies were detected in the sera of these patients against any of the investigated antigens.

Discussion

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References

We report eight patients with sensitization to bird serum proteins who developed allergic respiratory symptoms upon exposure to these antigens by inhalation. They also experienced allergic symptoms after the ingestion of eggs. Immunologic studies showed that the symptoms were due to IgE-mediated sensitization to allergens present in egg yolk and avian extracts. All patients had positive skin tests to egg yolk, chicken serum, chicken meat, chicken feathers, and CSA. On the basis of skin tests, specific IgE and immunoblot results, we identified CSA as the main allergen causing bird-egg sensitization in these patients. We further demonstrated by means of specific bronchial provocation and oral challenges that CSA may act both as an inhalant and a food allergen.

De Maat-Bleeker et al. (1) first reported the association of hypersensitivity to ingested egg yolk with rhinitis and asthma caused by exposure to a parrot in an older woman. By RAST inhibition, Mandallaz et al. (3) demonstrated that livetin, the water-soluble fraction of egg-yolk proteins, was the major cross-reacting antigen found in bird feathers and egg yolk, and they coined the term “bird-egg syndrome” to designate this IgE-mediated association of inhalant and food allergy.

There is extensive cross-antigenicity among the serum proteins of various avian species, particularly among the albumins and β-glycoproteins (27). Egg yolk contains significant quantities of serum proteins, since livetins are derived from the blood of the hen (28). Williams (29) identified α-livetin as chicken serum albumin, a protein of 65–70 kDa. Szépfalusi et al. (5) pointed out that CSA (α-livetin) is a cross-reactive allergen in the bird-egg syndrome. The results of immunoblot and inhibition assays in our study are in agreement with previous reports and show that CSA is a cross-reactive allergen present in egg yolk and chicken serum and feathers. However, the clinical relevance of sensitization to CSA in this syndrome had not been fully documented.

In 1998–9, 62 patients were diagnosed with egg allergy (mainly to egg white) in our allergy clinic; eight of these individuals (12.9%) presented a combined sensitization to egg yolk and bird feathers. Although it has been reported that patients with bird-egg syndrome are mainly women (5), there was a majority of male patients in the present study.

According to the sequence of appearance of symptoms, the route of sensitization in bird-egg syndrome seems to be primarily respiratory, with ensuing food allergy to egg by cross-sensitization. Nevertheless, a previous sensitization to egg-yolk proteins could also predispose some patients to respiratory symptoms from birds (30).

Allergy to hen's egg white usually occurs in atopic children, who tend to outgrow this particular food allergy. Moreover, several egg-white allergens have been incriminated in asthma due to the inhalation of egg-white aerosols (13–18), which generally affects adults exposed in the workplace. The major allergens from egg white are ovomucoid (Gal d 1), ovalbumin (Gal d 2), conalbumin (Gal d 3), and lysozyme (Gal d 4) (31). In this paper, we have confirmed the important role of CSA as an inhalant and food allergen, and we propose the name Gal d 5 for this protein (accession no. X60688). The partial cross-reactivity between CSA and conalbumin indicate the presence of common epitopes in these proteins, and this could explain why some patients with bird-egg syndrome have positive skin or serologic tests to conalbumin and egg white. Nevertheless, a concomitant sensitization to both egg-yolk and egg-white proteins cannot be ruled out.

The number of domestic birds in the USA and Germany has been estimated at 25–30 million (32) and over 8 million (33), respectively. An epidemiologic study carried out in Spain showed that 11% of 4000 patients attending an allergy clinic for the first time kept birds at home (34). The birds most commonly kept as pets are budgerigars, parrots, and canaries. Exposure to birds may cause allergic symptoms such as rhinoconjunctivitis, nocturnal cough, and asthma. It has been suggested that pet birds may be an allergenic problem as great as that of cats and dogs (32). The analysis of the indoor factors associated with asthma symptoms in Austrian children aged 6–9 years showed that keeping a bird as a domestic pet was associated with a significantly increased risk of childhood wheezing (35).

Birds regularly release feather particles that contaminate the ambient air of the household with microscopic inhalant particles (32, 33). The specialized feathers (pulviplumes or powder down) of many birds, particularly those of the Psittacidae family (e.g., budgerigars, parrots, parakeets, and cockatoos), are coated with a very fine dust resembling talcum powder (33). The powder particles, which are granular, rod-shaped, or splinter-shaped, are about 1 µm in diameter and can be easily inhaled and deposited in the airways (32, 33). All domestic and most wild birds have uropygial or preen glands, which secrete a lipid-proteinaceous substance. The bird constantly dips its beak into this gland, cleaning and waterproofing the feathers. The sebaceous material and saliva dry on the feathers along with the protective, powdery substances, thus creating more airborne inhalant allergens (32). The growth and shedding of cornified epithelium of the bird skin is an added factor in the production of airborne allergens. Bird antigen can be detected in the home environment for prolonged periods of time after bird removal and cleanup (36). It is possible that nondomestic birds and free-roaming city pigeons may also cause asthma, which may be misdiagnosed.

However, there are a few studies about the allergens in feathers that may cause IgE-mediated allergic sensitization and asthma. Tauer-Reich et al. (33) identified several allergenic components in the feather extracts and serum proteins of budgerigar, parrot, pigeon, canary, and hen, with masses of 20–30 kDa and 67 kDa, the last probably corresponding to CSA.

By using air sampling and immunoblotting, we have demonstrated that CSA is recoverable from the air of a house with 11 free-flying budgerigars. Thus, CSA can be detected in the domestic environment of homes with bird pets, indicating that CSA may act as an aeroallergen able to elicit respiratory symptoms. Although the concentration of airborne CSA that causes sensitization or provokes asthma symptoms remains undetermined, concentrations of CSA of 3–34 µg/ml were sufficient to induce asthmatic responses in our patients in bronchial provocation tests.

The allergenicity of serum albumins is well known, and the involvement of these proteins in allergy to pet dander and in hypersensitivity to milk and bovine meat has been previously reported (37). Despite a high degree of sequence homologies among albumins from different animal species, a remarkable variability of IgE cross-reactivities has been observed among patients allergic to animals (38). The antigenicity of albumins can be reduced by heat treatment, possibly as a consequence of the structural modifications originated by the heating process (39, 40). Werfel et al. (40) showed that bovine serum albumin (BSA) is one of the heat-labile proteins in beef extract, and suggested that sensitization to these heat-labile proteins might explain why some patients tolerate well-cooked but not medium-rare beef. However, Fiocchi et al. (41) found that heating BSA at 100°C for 5 min did not significantly reduce the allergenicity of this protein. Our results demonstrate that heating reduces, but does not abolish, the allergenicity of CSA and livetins, and this fact could also explain why some sensitized patients may tolerate well-cooked, but not raw, eggs (42).

We conclude that CSA can act as both an inhalant and a food allergen in patients with the bird-egg syndrome. In view of the clinical and immunologic evidence that CSA is an egg-yolk/bird allergen implicated in the bird-egg syndrome, we propose that this allergen be designated Gal d 5 according to the International Union of Immunological Societies nomenclature (43).

Acknowledgment

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References

We thank Dr Ana Jiménez for help in air sampling.

References

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgment
  7. References
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