Parvalbumin in fish skin-derived gelatin: is there a risk for fish allergic consumers?

The major allergen parvalbumin was purified from cod muscle tissues, and polyclonal antibodies were raised towards it. The antibodies were tested for specificity and an enzyme-linked immunosorbent assay (ELISA) was developed using these antibodies. The ELISA was applied to measure parvalbumin in cod skin, the starting material for fish gelatin made from deep sea, wild fish. The ELISA was sufficiently sensitive (LLOQ = 0.8 ng ml(-1) in extracts, corresponding to 0.02 µg of parvalbumin per g of tissue), and did not cross-react with common food constituents. Fish gelatin, wine and beer, matrices for the potential use of this ELISA, did not cause disturbance of the assay performance. The data show that the parvalbumin content in cod muscle tissue is 6.25 mg g(-1), while the skins contained considerably less, 0.4 mg g(-1). Washing of the skins, a common industrial procedure during the manufacturing of fish gelatin, reduced the level of parvalbumin about 1000-fold to 0.5 µg g(-1), or 0.5 ppm. From 95 commercial lots of fish gelatin it is shown that 73 are below 0.02 µg g(-1) parvalbumin. From the other 22 lots, the one with the highest concentration contained 0.15 µg g(-1) of parvalbumin. These levels are generally assumed to be safe for fish-allergic individuals.

Presentation of allergen in different food preparations affects the nature of the allergic reaction--a case series.

BACKGROUND: Characterization of fatal and non-fatal reactions to food indicates that the majority of reactions are due to the ingestion of prepared foods rather than the non-processed allergen. In an ongoing study that used a double-blind placebo-controlled food challenge to investigate peanut allergy and clinical symptoms, the observed reaction severity in four of the first six subjects was greater than anticipated. We hypothesized that this was due to differences in the composition of the challenge vehicle. OBJECTIVE: The aim was to investigate whether the severity of observed challenge reactions would be repeated on re-challenge with a lower fat challenge vehicle. METHODS: Peanut-allergic subjects were re-challenged with a lower fat recipe after reacting more severely than was anticipated to an initial peanut challenge. Similar challenge vehicle recipes were used, the only difference being the lower fat content (22.9% compared with 31.5%). The peanut content of the two recipes was analysed using RAST inhibition studies and ELISA tests. RESULTS: Three of four subjects reacted to much smaller doses of peanut protein on re-challenge (mean dose equivalence - 23 times less peanut) with the lower fat recipe. RAST inhibition showed that neither recipe altered epitope recognition. The higher fat recipe required twice as much peanut to cause 50% inhibition. ELISA detected far lower levels of peanut in the higher fat recipe (220 000 parts per million (p.p.m.)) than in the lower fat recipe (990 000 p.p.m.). CONCLUSION: The fat content of a challenge vehicle has a profound effect on the reaction experienced after allergen ingestion. This is another factor to be considered in assessing the risk of certain foods to food-allergic consumers and adds another dimension to clinical, research and regulatory practice.

An evaluation of the sensitivity of subjects with peanut allergy to very low doses of peanut protein: a randomized, double-blind, placebo-controlled food challenge study.

BACKGROUND: The minimum dose of food protein to which subjects with food allergy have reacted in double-blind, placebo-controlled food challenges is between 50 and 100 mg. However, subjects with peanut allergy often report severe reactions after minimal contact with peanuts, even through intact skin. OBJECTIVE: We sought to determine whether adults previously proven by challenge to be allergic to peanut react to very low doses of peanut protein. METHODS: We used a randomized, double-blind, placebo-controlled food challenge of 14 subjects allergic to peanuts with doses of peanut ranging from 10 microg to 50 mg, administered in the form of a commercially available peanut flour. RESULTS: One subject had a systemic reaction to 5 mg of peanut protein, and two subjects had mild objective reactions to 2 mg and 50 mg of peanut protein, respectively. Five subjects had mild subjective reactions (1 to 5 mg and 4 to 50 mg). All subjects with convincing objective reactions had short-lived subjective reactions to preceding doses, as low as 100 microg in two cases. Five subjects did not react to any dose up to 50 mg. CONCLUSION: Even in a group of well-characterized, highly sensitive subjects with peanut allergy, the threshold dose of peanut protein varies. As little as 100 microg of peanut protein provokes symptoms in some subjects with peanut allergy.

Identification of a Brazil-nut allergen in transgenic soybeans.

BACKGROUND: The nutritional quality of soybeans (Glycine max) is compromised by a relative deficiency of methionine in the protein fraction of the seeds. To improve the nutritional quality, methionine-rich 2S albumin from the Brazil nut (Betholletia excelsa) has been introduced into transgenic soybeans. Since the Brazil nut is a known allergenic food, we assessed the allergenicity of the 2S albumin. METHODS: The ability of proteins in transgenic and non-transgenic soybeans, Brazil nuts, and purified 2S albumin to bind to IgE in serum from subjects allergic to Brazil nuts was determined by radioallergosorbent tests (4 subjects) and sodium dodecyl sulfate-polyacrylamide-gel electrophoresis (9 subjects) with immunoblotting and autoradiography. Three subjects also underwent skin-prick testing with extracts of soybean, transgenic soybean, and Brazil nut. RESULTS: On radioallergosorbent testing of pooled serum from four subjects allergic to Brazil nuts, protein extracts of transgenic soybean inhibited binding of IgE to Brazil-nut proteins. On immunoblotting, serum IgE from eight of nine subjects bound to purified 2S albumin from the Brazil nut and the transgenic soybean. On skin-prick testing, three subjects had positive reactions to extracts of Brazil nut and transgenic soybean and negative reactions to soybean extract. CONCLUSIONS: The 2S albumin is probably a major Brazil-nut allergen, and the transgenic soybeans analyzed in this study contain this protein. Our study show that an allergen from a food known to be allergenic can be transferred into another food by genetic engineering.

Peanut sensitivity.

Peanuts are one of the most allergenic foods. The allergic reactions may vary in severity from mild urticaria to severe anaphylactic episodes and death. The prevalence of peanut sensitivity is unknown, but it may affect as many as 10% of allergic individuals. The chemistry of peanut proteins has been extensively studied. Two major protein fractions have been prepared from saline extracts of peanut flour, arachin and conarachin. A major peanut allergen termed "Peanut-1" has been isolated. However, a number of protein constituents, including the arachin and conarachin fractions, have been shown to be allergenic. The ability to diagnose peanut sensitivity accurately has been hampered by the lack of standardized peanut extracts. However, efforts are under way to prepare such standardized reagents. Treatment consists of avoiding peanut protein products and using self-administered epinephrine. A number of peanut protein-containing products are allergenic, although peanut oil is not. The peanut-allergic consumer should be instructed to carefully read labels of foods. This can at times, however, be misleading, because certain foods may be inadvertently contaminated by peanut proteins.

Histamine poisoning (scombroid fish poisoning): an allergy-like intoxication.

Histamine poisoning results from the consumption of foods, typically certain types of fish and cheeses, that contain unusually high levels of histamine. Spoiled fish of the families, Scombridae and Scomberesocidae (e.g. tuna, mackerel, bonito), are commonly implicated in incidents of histamine poisoning, which leads to the common usage of the term, "scombroid fish poisoning", to describe this illness. However, certain non-scombroid fish, most notably mahi-mahi, bluefish, and sardines, when spoiled are also commonly implicated in histamine poisoning. Also, on rare occasions, cheeses especially Swiss cheese, can be implicated in histamine poisoning. The symptoms of histamine poisoning generally resemble the symptoms encountered with IgE-mediated food allergies. The symptoms include nausea, vomiting, diarrhea, an oral burning sensation or peppery taste, hives, itching, red rash, and hypotension. The onset of the symptoms usually occurs within a few minutes after ingestion of the implicated food, and the duration of symptoms ranges from a few hours to 24 h. Antihistamines can be used effectively to treat this intoxication. Histamine is formed in foods by certain bacteria that are able to decarboxylate the amino acid, histidine. However, foods containing unusually high levels of histamine may not appear to be outwardly spoiled. Foods with histamine concentrations exceeding 50 mg per 100 g of food are generally considered to be hazardous. Histamine formation in fish can be prevented by proper handling and refrigerated storage while the control of histamine formation in cheese seems dependent on insuring that histamine-producing bacteria are not present in significant numbers in the raw milk.

Sensitivity to sulfited foods among sulfite-sensitive subjects with asthma.

Eight individuals with asthma who had been diagnosed as sulfite sensitive on the basis of double-blind capsule-beverage challenges were subjected to challenges with various sulfited foods, including lettuce, shrimp, dried apricots, white grape juice, dehydrated potatoes (as mashed potatoes), and mushrooms. Four of these patients failed to respond to challenges with any of the sulfited foods. The other four patients experienced a decrease in pulmonary function on double-blind challenges with sulfited lettuce. Two of three of these patients reacted to challenges with dried apricots and white grape juice; the fourth patient has not yet been challenged with these products. Only one of these four patients reacted to challenges with dehydrated potatoes and mushrooms, and, in this case, the response to double-blind challenges with dehydrated potatoes was not consistent. None of the sulfite-sensitive subjects with asthma responded to challenges with sulfited shrimp. It is concluded that sulfite-sensitive subjects with asthma will not necessarily react after ingestion of sulfited foods. The likelihood of a reaction is dependent on the nature of the food, the level of residual sulfite, the sensitivity of the patient, and perhaps on the form of residual sulfite and the mechanism of the sulfite-induced reaction.

False positive and false negative reactions encountered in the use of sulfite test strips for the detection of sulfite-treated foods.

The reliability of a qualitative test for sulfites in foods (Sulfitest sulfite test strips) was evaluated by comparison of the results obtained from the analysis of 90 food and beverage samples to results obtained by the quantitative, modified Monier-Williams method (the preferred procedure for sulfite analysis in foods of the Food and Drug Administration). The results obtained with the sulfite test strips compared favorably to the results of the Monier-Williams procedure when the sulfite test strips were used on sulfite-treated lettuce and raw or cooked potatoes. However, the strips yielded many false negative and false positive results with other types of foods. False positive results (strips indicated a substantial amount of sulfite when sulfite was not detectable by the Monier-Williams method) were obtained with fin fish, red meats, and poultry. False negative results (strips indicated the absence of sulfites when sulfite was detected at levels greater than 10 ppm total sulfur dioxide by the Monier-Williams method) were obtained with dried fruits and wines under certain conditions of testing. The false negative responses with the test strips may result in the hazardous consumption of foods with high levels of sulfites, such as dried fruit or wine, by a sulfite-sensitive individual. The false positive responses would not be hazardous but could lead sensitive individuals to avoid foods that could be safely consumed. Although the strips may be useful for the detection of sulfites in certain foods, such as lettuce and potatoes, their use by sulfite-sensitive individuals cannot be recommended because of the confusion and potential hazards posed by the false negative and false positive responses.

Prevalence of sensitivity to sulfiting agents in asthmatic patients.

Ingestion of sulfiting agents can induce wheezing in some asthmatic patients. However, neither the prevalence of sulfite sensitivity nor the clinical characteristics of the affected asthmatic population are known. In a prospective single-blind screening study, 120 non-steroid-dependent and 83 steroid-dependent asthmatic patients underwent challenge with oral capsules of potassium metabisulfite. Five non-steroid-dependent and 16 steroid-dependent asthmatic patients experienced a greater than 20 percent reduction in their one-second forced expiratory volume within 30 minutes following the oral challenge. Twelve of these sulfite reactors were rechallenged with metabisulfite capsules in a double-blind protocol. Under these conditions, only three of seven steroid-dependent patients had a positive response. Moreover, only one of five non-steroid-dependent patients had a response to double-blind challenge. On the basis of this challenge study, the best estimate of the prevalence of sulfite sensitivity in the asthmatic patients studied is 3.9 percent. This population, however, contained a larger number of steroid-dependent asthmatic patients than would be found in the general asthmatic population. It is concluded, therefore, that the prevalence of sulfite sensitivity in the asthmatic population as a whole would be less than 3.9 percent and that steroid-dependent asthmatic patients are most at risk.

Soybean oil is not allergenic to soybean-sensitive individuals.

We have previously demonstrated that peanut oil is not allergenic to peanut-sensitive individuals. Seven soybean-sensitive patients were enrolled in a double-blind crossover study to determine whether ingestion of soybean oil can induce adverse reactions in such patients. All subjects had histories of systemic allergic reactions (urticaria, angioedema, wheezing, dyspnea, and/or vomiting) after soybean ingestion and had positive puncture skin tests with a 1:20 w/v glycerinated-saline whole soybean extract. Sera from six of the seven subjects were tested by RAST assay for the presence of specific IgE antibodies to soybean allergens. All patients had elevated levels of serum IgE antibodies to the crude soybean extract; binding values ranged from 2.3 to 28.1 times that of a negative control serum. Before the oral challenges, all patients demonstrated negative puncture skin tests to three commercially available soybean oils and to olive oil (control). On four separate days, patients were challenged with the individual soybean oils and olive oil in random sequence. At 30-minute intervals, under constant observation, patients ingested 2, 5, and 8 ml of one of the soybean oils or olive oil contained in 1 ml capsules. No untoward reactions were observed with either the commercially available soybean oils or olive oil. Soybean oil ingestion does not appear to pose a risk to soybean-sensitive individuals.

Allergenicity of various peanut products as determined by RAST inhibition.

Extracts of 19 different peanut products and peanut oil were tested for their allergenicity by the radioallergosorbent test inhibition assay using a crude peanut extract from raw peanuts as the standard for comparison. Seventeen of the extracts were able to competitively inhibit the binding of serum IgE from peanut-sensitive patients with the solid-phase raw peanut extract. Peanut oil and the extract from hydrolyzed peanut protein did not inhibit binding, which suggests that these products are not allergenic. The peanut hull flour extract showed a slight ability to inhibit binding, suggesting that this product contains minor amounts of the peanut allergen.