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Rapid production of recombinant allergens in Nicotiana benthamiana and their impact on diagnosis and therapy.

BACKGROUND: Type I allergies are immunological disorders that afflict a quarter of the world's population. Recombinant allergens have improved the diagnosis of allergic diseases and allow the formulation of new therapeutic approaches. Over 50% of all allergens are of plant origin. OBJECTIVE: We have applied a novel method of overexpressing plant allergens in the tobacco-related species Nicotiana benthamiana. METHOD: This method is based on the use of a chimeric tobacco mosaic virus that harbors a foreign gene sequence and directs its transcription after the infection of the host plant. RESULTS: We have expressed the model allergen Bet v 1, the major birch pollen allergen, and two Hevea brasiliensis latex allergens, the spina-bifida-associated allergens Hev b 1 and Hev b 3, in N. benthamiana using such a viral vector. Bet v 1, Hev b 1 and Hev b 3 produced by this method were recognized by patients' IgE suggesting that the plant-produced allergens were properly folded. Nonpurified Bet v 1 expressed in N. benthamiana leaves had the same immunogenicity as purified Bet v 1 expressed in Escherichia coli or natural Bet v 1 when tested in a murine model of type I allergy. CONCLUSION: We conclude that this plant expression system offers a viable alternative to fermentation-based production of allergens in bacteria or yeasts.

Allergens↗

Urticaria and rhinitis to shrubs of Ficus benjamina and breadfruit in a banana-allergic road worker: evidence for a cross-sensitization between Moracea, banana and latex.

BACKGROUND: We report the case of a road worker with a food allergy to banana, who developed urticaria and rhinitis when cutting shrubs of Ficus benjamina and breadfruit. He did not develop an allergy to latex of Hevea brasiliensis. RESULTS: Sensitization to latex of F. benjamina, H. brasiliensis, breadfruit and banana was demonstrated using skin tests and specific IgE measurements. RAST inhibitions procedures showed that specific IgE to breadfruit latex cross-reacted more strongly with latex of H. brasiliensis and banana than with latex of F. benjamina with the same extract. CONCLUSION: Given the wide distribution of Moracea trees in tropical regions, sensitization to latex of H. brasiliensis and banana could be a consequence of sensitization to Moracea members; F. benjamina does not seem to be the only Moracea responsible for cross-allergy with latex and fruit. Consequently, it seems interesting to test other members of the Moracea family in patients sensitized to latex of H. brasiliensis and banana. Sensitization to breadfruit could be a risk factor for sensitization to latex of H. brasiliensis.

Adult↗

Oral allergy syndrome to fig.

BACKGROUND: The few cases of food allergy to fig reported to date, whose main manifestations were anaphylactic reactions, have been related to a cross-sensitisation to weeping fig (Ficus benjamina) or to the 'latex-fruit syndrome'. Here we report on two cases of the oral allergy syndrome (OAS) to fig in patients whose main allergic manifestations were related to sensitisation to grass and birch pollens. METHODS: The patients were characterised by clinical history, skin prick tests (SPT) with commercial and in-house extracts, prick-by-prick test, specific IgE measurements and challenge tests. PBS-soluble and insoluble extracts of both fig skin and pulp were examined for the presence of potential allergens by IgE immunoblotting. RESULTS: Both patients showed OAS followed by respiratory symptoms when challenged with fig. They were negative in both specific IgE detection and SPT with commercial extracts of fig and many other plant materials, including F. benjamina and Hevea Brasiliensis, while grass and birch pollens gave positive results. Prick-by-prick tests and SPT with in-house extracts indicated that the fig skin had a much higher allergenicity than the pulp. Despite negative IgE detection by the CAP assay, immunoblotting experiments showed that potential fig allergens were PBS-soluble and present only in the skin of the fruit. CONCLUSIONS: OAS to fig followed by respiratory symptoms can be present in patients not sensitised to weeping fig or having the latex-fruit syndrome. Different parts of the fig can have different allergenicities, the most important allergens being proteins related to the skin of the fruit. Improved commercial fig extracts to be used for the diagnosis of this type of allergy have to be developed.

Adult↗

Identification of hevamine and hev B 1 as major latex allergens in Taiwan.

BACKGROUND: Proteins from latex gloves have been documented to trigger occupational latex allergy among health care workers. Allergen characterization of latex glove extract has never been studied in Taiwan. This study aimed to identify allergenic proteins from latex gloves. METHODS: Crude extracts of latex gloves were prepared with phosphate-buffered saline and 20 medical workers with a history of latex allergy were enrolled in this study. The specific IgE antibody was determined by the Pharmacia CAP system and in-house enzyme-linked immunoassay and immunoblotting. The target proteins were excised from two-dimensional PAGE and analyzed by electrospray ionization tandem mass spectrometry. RESULTS: Immunoblotting of glove extracts revealed three IgE-binding proteins at a molecular mass of 45, 30 and 14 kDa. Peptide mass fingerprinting revealed that the protein at 45 kDa, which was recognized by 10% (2/20) of atopic sera tested, was an allergenic lipolytic esterase from Hevea brasiliensis (Hev b 13). The 30- and 14-kDa proteins, which were recognized by 55% (11/20) and 85% (17/20) of patients' sera, were found to be hevamine and rubber elongation factor (Hev b 1), respectively. CONCLUSIONS: Our results indicated that hevamine and Hev b 1 are the major allergens from latex gloves in Taiwan, which differs from the reports in Western countries.

Adult↗

[Richard Spruce, botanist-South America's explorer].

Between 1849 and 1864, the English botanist and explorer Richard Spruce carried out a detailed study of the Amazon flora and the costumes of the peoples who inhabited the region. To date a large part of the existing knowledge about several botanical families in the region stems from this scientist's efforts. His comprehensive interests, his detailed and precise descriptions were outstanding: nothing seems to have been left out of his scrutiny and recording aptitude. Not only was Spruce a remarkable botanist but he was also a distinctive anthropologist, linguist (he knew French, Spanish and Portuguese), geologist and geographer, as well as an acute sociological observer of the political systems and habits of the Amazonian and Andean trips in which has has been. He could thus make a considerable contribution to the understanding of indigenous beliefs and practices, as well as to the knowledge and uses of plants within the Amazonian context. Also important was his participation in the economic exploration of local species, particularly as regards the Hevea and the Cinchona genera.

Anthropology↗

Biocompatibility of natural latex implanted into dental alveolus of rats.

The present study investigated the biocompatibility of a biopolymer based on vegetable latex extracted from the Hevea brasiliensis rubber tree, implanted into the bony alveolar cavity after dental extraction in rats. A granule of latex (area = 0.25 +/- 0.04 mm(2)) was implanted inside the alveolus immediately after extraction of the upper right incisor, and the animals were sacrificed 7, 21 and 42 days after the procedure. The hemi-maxillas were decalcified and processed for embedding in paraffin to obtain semi-serial longitudinal sections 5 mum thick, and then stained with hematoxylin-eosin. The latex granule was observed in the cervical third of the alveolus without any foreign body reaction, or persistence of the initial acute inflammatory reaction. Bone repair in the areas adjacent to the material was quantified, and a decrease was noted in the thickness of the fibrous capsule surrounding the implants from 92.8 +/- 9.3 microm on day 7 to 9.4 +/- 1.8 microm on day 42 (ANOVA, P = 0.01). The quantitative data confirmed acceleration of bone formation (statistically significant at 5%) in parallel with a decrease of connective tissue in the areas around the implants. These results show that the tested material is biologically compatible, and progressively integrated into the alveolar bone, simultaneously accelerating bone formation and playing an important role in the healing process.

Absorbable Implants↗

Characterization of latex antigen and demonstration of latex-specific antibodies by enzyme-linked immunosorbent assay in patients with latex hypersensitivity.

Two latex antigens, one extracted from surgical gloves (GE) and the other from the sap of Hevea brasiliensis plant (RPE) were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional gel electrophoresis. These antigens were used to detect latex specific IgE and IgG antibodies by enzyme-linked immunosorbent assay (ELISA) in the sera of patients with meningomyelocele (spina bifida) and of normal controls. Of the 36 patients studied, 26 had wheal-and-flare skin-prick test reactivity to latex antigen with 11/26 having had a history of anaphylaxis to latex products. Twenty three of the 26 sera from skin-test positive patients and 10/11 patients with history of anaphylaxis demonstrated significant levels of latex specific IgE and IgG in the sera, whereas only 1/14 normals showed significant antibodies to latex. The remaining 10 patients, all skin-test negative with latex antigens, showed only low levels of antibodies. The findings indicate that the ELISA used in the present study employing partially characterized antigens has sensitivity and specificity to detect latex specific antibodies in the sera of suspected patients and can be used for presumptive diagnosis of latex allergy.

Allergens↗

Allergy to natural rubber latex: a growing problem.

Immediate, IgE-mediated allergy to natural rubber latex has recently been found to be responsible for contact and generalized urticaria, protein contact dermatitis, rhinitis, asthma and even fatal anaphylactic reactions. The proteins present in latex from the rubber tree (Hevea brasiliensis) can still persist in rubber end-products where they are capable of sensitizing exposed users. Atopic people and those with hand eczema are prone to become sensitized by natural rubber latex, which often causes occupational problems in connection with the use of protective gloves. But people blowing up balloons and those using condoms are also at risk if their symptoms are misinterpreted. Severe adverse reactions caused by rubber products in medicare cannot be prevented if the diagnosis has not been made before operations and medical examinations. So the most important task today is to develop reliable and safe methods for diagnosing natural rubber latex allergy and to manufacture rubber products with fewer allergens.

Gloves, Protective↗

Characterization of T cell responses to Hev b 3, an allergen associated with latex allergy in spina bifida patients.

The prevalence of type I allergy to Hevea brasiliensis latex is particularly high among individuals with frequent exposure such as health care workers and patients with spina bifida (SB). Due to a birth defect of the spinal canal and the resulting neurological and orthopedic defects, these patients require multiple surgeries during childhood. SB patients display a unique pattern of sensitization: IgE-reactivity is preferentially directed against Hev b 3 and Hev b 1, two latex allergens with high sequence similarity. In this study, we analyzed the T cell response to Hev b 3 in latex-allergic SB patients using poly-, oligo-, and monoclonal T lymphocyte cultures. All T cell clones (TCC) were CD3/CD4-positive and expressed the alphabeta TCR. According to their cytokine production pattern (IL-4 vs IFN-gamma), 12 of 21 TCC were classified as Th2-like, 2 of 21 were Th1-like, and 7 of 21 belonged to a Th0-like subset. Using 11 T cell lines and 21 TCC, nine T cell stimulating fragments were determined out of 52 overlapping 12-mer peptides representing the complete amino acid sequence of Hev b 3. Ag presentation of one dominant T cell epitope could be associated with a four-amino acid binding motif (YSTS, position 11-13) in the beta 1 chain of HLA-DR molecules expressed by the respective patients. No reactivity was observed when Hev b 3-reactive T cell lines or TCC were incubated with peptides representing homologous parts of the Hev b 1 molecule, i.e., no cross-reactivity between Hev b 3 and Hev b 1 at the T cell level was evident.

Allergens↗

[Latex: a complex allergy].

The authors review several of the most important aspects of latex allergy, an increasing problem in Public Health, which should be understood by all health professionals. After briefly presenting the history of the origin latex, from Hevea brasiliensis the authors describe the antigens of latex: Hev b1 to Hev b8, major allergens. They also note the crossed reactivity not only with foods, exotic fruits, but also with pneumoallergens and in particular the pollens. The groups at risk are essentially workers in the latex industry, health professionals and finally infants with spina bifida or other severe urological anomaly. The clinical signs are reactions of type 1 hypersensitivity, to urticaria and/or angio-oedema and anaphylactic shock. Diagnosis is based on a search for specific serum IgE, skin tests and provocation tests. Prophylaxis depends on removal of all substances that are based on latex, especially replacement of gloves with vinyl, but also on a food diet that excludes all foods that have a cross-reactivity with latex.

Allergens↗

[Latex allergy. An not rare intraoperative event].

Latex allergy has become a real problem among both surgical staff (paramedics and physicians) and patients especially pediatric patients with urogenital malformations and spina bifida. Latex allergy is produced from both natural molecules which compose the substance produced from Hevea brasiliensis (rubber tree) and industrial additives contents in latex devices. Diagnosis of latex allergy may be carried out through a preoperative Prik-test. A characteristic of latex allergy reaction is the starting of symptoms (more than 15 minutes after allergen contact). Pathophysiology of latex allergy is the same of all allergic reactions; it is an antigen-antibody reaction and type I or II reaction may occur. There are a lot of devices that surgeons and anesthesists use in the operative theatre and that should not be used in presence of a patient with latex allergy. Guaranteed latex-free devices should always be present in store.

Humans↗

Prevalence of allergy to latex in the pediatric population.

Latex is a substance that is extracted from the plant Hevea Brasiliensis, and world production is 6 million metric tons per year. After gathering it undergoes a series of processes in which ammonia, vulcanization-accelerating additives or anti-oxidants are added. All of this may constitute the antigenic load that latex contains. It is used in medicine for gloves, drainage tubes, dental implants and a growing number of products from condoms to sport products or automobile components. Over fifteen different allergenic bands have been described, with molecular weights of between 2 and 100 kDa. In a joint study, USA-Finland identified three antigenic bands that predominate according to the population type studied (2). In children with spina bifida, congenital urogenital abnormalities or those submitted to multiple surgical interventions, the band of 27 kDa is predominant. This antigen has not been detected in adult serum, which suggests that contact with the antigen is through the mucous membranes and the sensitization is triggered from here. The incidence of allergy to latex in the general population is not known, but it seems to be lower than 1%. Turjanmaa (1) establishes a frequency of 0.125% (1/800) in patients submitted to general surgery. The incidence of sensitization to latex varies according to the population studied. Among the population considered to be at risk are the workers of the health environment, where the incidence is between 2.6 and 16.9%, whereas in the general population the percentage is around 1%. Sensitivity to latex is of great importance in patients suffering from spina bifida, in whom an incidence of between 28 and 67% has been found. We present our experience in a group of children attending our service for the first time for diverse reasons of supposed allergic etiology. The objective is to determine the incidence of sensitization to latex according to the diagnostic methodology [cutaneous test or by determination of specific IgE (CAP)]; according to the type of patient (atopic or non-atopic), the direct relationship with latex material and the role that can be played by a history of surgical intervention. In our experience with 282 children studied in our pediatric allergology service using diverse methods, the incidence of allergy to latex is 3.19%. Nevertheless, if we analyze this percentage we observe that if the diagnosis is based exclusively on cutaneous tests, it is only 1.08%; if to establish a diagnosis we used exclusively the determination of specific IgE (CAP) we would label 7. 2% of our children as allergic. Atopy is a factor that facilitates sensitization. In our sample, the incidence among the atopic population is 4.4%, though this percentage may vary between 1.69% and 9.5% depending on the methodology used. There are several hypotheses for explaining these discrepancies in the diagnostic tests. Although there do not seem to be differences regarding the ammonia content of the different lots, it seems that the differential factor could lie in the type of extract and in whether it is commercial or a natural latex extract. It could be a consequence of the existence of a prophyllin, so it has not been ruled out that a part of the IgE is an antiprophyllin. The presence of different epitopes would mean that each of them has the ability to produce its own specific IgE, though the RAST/CAP was not able to differentiate them and identified them as whole. The rate of allergy to latex in a group of children suffering from myelomeningocele is 80%. At a paediatric level, sensitization to latex is influenced by the means of contact, the duration of the exposure to the antigen and the fact that the exposure occurs early, which in the case of children with myelomeningocele is associated with the immaturity of the defence mechanism of the mucous membranes.

Child↗

A new quantitative in vitro for the detection of latex-specific IgE antibodies.

Immediate-type hypersensitivity to latex allergens has resulted in anaphylactic shock and death in numerous reported cases. The allergenic proteins of latex are contained within the natural rubber extract of Hevea brasiliensis and are eluted into the final product during the manufacturing process. The quantity and types of latex allergens found in different latex products depends on the manufacturing process. Not all of these allergens are available for use in the latex prick skin test, and as a result, such tests may not be conclusive. Furthermore, application of such allergens to the skin of undiagnosed hypersensitive individuals may have harmful effects on their health. Therefore, it is important to be able to utilize in vitro methods, which reliably identify latex allergy without placing hypersensitive individuals at risk. We have developed a relatively simple and new enzyme immuno-assay (EIA) method for the detection of latex allergy. This in vitro method is quantitative and allows for the classification of allergy to latex in a short time. In comparative studies, ninety-nine serum specimens with documented clinical history of latex allergy were tested by this method, and the results paralleled those of the skin prick test performed by an independent group. The data showed that the specificity and sensitivity of our assay approaches 97.5% and 100%, respectively. We conclude that, by using a simple assay, the detection of specific IgE to latex proteins may be valuable for screening individuals and for the diagnosis of allergy to latex.

Antibody Specificity↗

Couriers of asthma: antigenic proteins in natural rubber latex.

Natural rubber latex (NRL) is a milky, white liquid containing the polymer cis-1,4-polyisoprene, derived from the laticifer cells of the rubber tree, Hevea brasiliensis. Reports of allergic reactions to NRL, ranging in severity from skin rashes to anaphylaxis and death, have been increasing. High rates of latex allergy are seen in children with spina bifida. Healthcare workers, especially those who frequently use powdered latex medical gloves, can develop NRL allergy and asthma, often resulting in considerable disability. Atopy and, possibly, pre-existing dermatitis predispose to sensitization and allergic symptoms. NRL protein antigens are found in many NRL products and also have been shown to bind to cornstarch glove-donning powders. Of the approximately 240 polypeptides in NRL, nearly 60 are antigenic, and nine have been identified and registered (Hev b 1&ndash9). Many natural latex antigens share epitopes with structural proteins and enzymes from other plant species. Current knowledge indicates that proposed reductions in total glove protein and limitations on the use of powder on NRL gloves are likely to result in a decline in the prevalence and severity of NRL allergies among healthcare workers.

Antigens↗

[Latex allergy and cross-reactions: a new threat?].

Allergy to latex proteins has become an important problem, especially in the medical profession, but also in people who have regularly contact with latex products. The responsible antigens are proteins, which are present in the natural rubber latex (NRL), extracted from Hevea brasiliensis. These proteins are important in the synthesis of rubber and in the defence of the plant against external noxes (pathogenesis related proteins). Life threathening reactions occur more frequently in the operating theatre. Sensitisation can occur via different routes (aerogen, skin, parenteral, ...) but aerogenic sensitisation seems to be very important. Starch particles are important vectors of the antigens. Diagnosis is made via history, confirmed by demonstration of specific IgE via skin test (in vivo) or via an in vitro method (e.g. CAP, ELISA). Immunological cross reactions are very frequent with other plant or fruit allergens but cross allergy is less frequent (sensitivity and specificity of the in vitro test varies considerably depending on the nature of the fruit or vegetable investigated). There is also a cross reactivity with Ficus. Prevention with latex allergens becomes primordial in hospital settings. The industry is aware of the problem and produces more and more synthetic latex products.

Cross Reactions↗

Allergy to Ficus benjamina.

The clinical history of 16 patients found to have specific IgE antibodies against Ficus benjamina was analyzed in terms of allergic symptoms, clinical and biological cross-sensitisation to other allergens and compared to the data found in the literature. Two different groups of patients were studied. Group A consisted of 13 patients who became sensitised through contact with ficus plants and experienced symptoms upon exposure. Their main symptoms where conjunctivitis, rhinitis, asthma, eyelid oedema or urticaria. Of these patients 11 had other atopic manifestations. Two persons had no other allergy. 10 patients experienced symptoms at home, 2 at the working place and 1 while being in a restaurant. One patient had a crossreactive food allergy to figs. Group B consisted of 3 highly atopic patients who had also a sensitization both to latex (Hevea brasiliensis) and to Ficus benjamina. They had no clinical history of allergic reactions provoked by ficus plants.

Adolescent↗

Latex allergy.

Natural rubber latex (NRL) has been exploited commercially for more than 100 years. It is derived from the milky sap of the Hevea brasiliensis tree, found in tropical climates such as Southeast Asia. Latex is harvested from the tree by a process called tapping, which has been described as a controlled wounding of the tree. The world's total consumption of natural rubber latex is about 5,5 million tons per year. The liquid which is collected from the tree is called field latex and is made up of water, 30%to 40% rubber, and about 3% nonrubber material.

Journal Article↗

Aeroallergens in clinical practice of allergy in India. An overview.

Allergic diseases such as bronchial asthma, allergic rhinitis and atopic dermatitis are dramatically increasing all over the world including developing countries like India. Today, more than 30 % of the population is known to suffer from one or other allergic ailment. Major causative agents implicated are pollen grains, fungal spores, dust mites, insect debris, animal epithelia, etc. Several aerobiological studies have been conducted in different parts of the country to ascertain aerial concentration and seasonality of pollen grains and fungi. Recently, an "All India Coordinated Project on Aeroallergens and Human Health" was undertaken to discover the quantitative and qualitative prevalence of aerosols at 18 different centres in the country. Predominant airborne pollen are Holoptelea, Poaceae, Asteraceae, Eucalyptus, Casuarina, Putanjiva, Cassia, Quercus, Cocos, Pinus, Cedrus, Ailanthus, Cheno/Amaranth, Cyperus, Argemone, Xanthium, Parthenium and others. Clinical and immunological evaluations have revealed allergenically important texa - some of them for the first time. Allergenically important pollen are Prosopis juliflora, Ricinus communis, Morus, Mallotus, Alnus, Querecus, Cedrus, Argemone, Amaranthus, Chenopodium, Holoptelea, Brassica, Cocos, Cannabis, Parthenium, Cassia and grasses. Further cross-reactivity of the IgE antibodies is a common phenomenon among various pollen allergens. Ricinus communis pollen from commonly growing weeds in India, cross-reacts with latex (Hevea brasiliensis), Mercurialis annua and also with seeds of Ricinus communis - all belonging to family Euphorbiaceae. Areca catechu cross-reacts with other members of Arecaceae such as Phoenix sylvestris, Cocos nucifera and Borassus flabelifer. Several reports on pollen and fruit syndrome have been analyzed. Experiments conducted by us revealed that pollutants (NO(2) and SO(2)) not only affect pollen morphology but also changes their allergenic potency. Immunotherapy with recombinant proteins having similar epitopes from different allergens have been advocated, besides allergen avoidance.

Air Pollutants↗