[Preliminary study on the enzymatic activity of the latex lutoids of Hevea brasiliensis. Distribution of acid phosphatase, beta-glucosidase and cathepsin in latex].
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Hevein, a lectin-like protein is the major protein of vacuolar structures called lutoids in the latex of rubber trees. We have shown both by in planta and ex planta studies that hevein is involved in the coagulation of latex by bringing together rubber particles. This polyvalent bridging between hevein and rubber particles is mediated by N-acetyl-D-glucosamine and involves a receptor glycoprotein of 22 kDa, which is localized on the surface of the rubber particles. The proposed role of hevein helps us to understand the mechanism of coagulation of latex and assigns a physiological intracellular function to one of the smallest lectins.
BACKGROUND: Ornamental Ficus benjamina (FB) has been recognized as a new indoor allergen. Little is known about the prevalence in moderately exposed subjects and the proposed association with fruit and Hevea latex hypersensitivity. OBJECTIVE: To study the prevalence of FB sensitization and the relationship with Hevea latex allergy, to identify cross-reacting fruits, and to characterize the responsible allergens. METHODS: A skin prick test solution prepared from FB latex (200 microg/mL) was included in our routine screening programme for suspect inhalant allergy. Patients reacting with the FB extract were further skin tested with exotic fruits by the prick-to-prick method. Inhibition of fig and FB CAP by FB latex, fig (Ficus carica), kiwi, the thiolproteases ficin and papain, Hevea latex and rHev b 6.02 (hevein) was performed in selected patients. RESULTS: Of 2662 patients with a positive skin test to any aeroallergen, 66 (2.5%) reacted with FB. Ten patients showed isolated sensitization to FB. Although FB-positive subjects were more often co-sensitized to Hevea latex than FB-negative (10.6% vs 3.8%, P< 0.01), nearly 90% tested negative for Hevea latex. Sensitization to FB was specifically associated with positive skin tests to fresh fig (83%), dried fig (37%), kiwi fruit (28%), papaya (22%), avocado (19%), banana (15%), and pineapple (10%) (n = 54). Clinical reactions were reported mainly from fresh and dried fig and kiwi (47%, 60%, and 64%, respectively, of skin test-positive patients), including seven patients with systemic reactions (urticaria, angiooedema, asthma). CAP to fig in 11 patients with clinical fruit allergy was inhibited on average by 87% by FB latex, 89% by fresh fig, 80% by dried fig, 38% by kiwi (100 microg/mL each), and by 59% and 44% by ficin and papain (50 microg/mL), respectively. No inhibition was obtained with Hevea latex and rHev b 6.02. CAP to FB was inhibited on average by 95% by FB, 60% by fresh fig, 41% by ficin, 29% by papain, and less than 7% by rubber latex allergens. CONCLUSIONS: Sensitization to FB latex is found in 2.5% of atopic individuals and mostly occurs independently of Hevea latex allergy. Sensitization is commonly associated with allergic reactions to figs and other tropical fruits ('Ficus-fruit syndrome'). This cross-reactivity is mediated at least in part by thiolproteases.
BACKGROUND: Association between allergy to Ficus benjamina and Hevea brasiliensis, two botanically unrelated plants, was suspected in consequence of two clinical observations. Symptoms were rhinitis and asthma. This study was undertaken to assess the in vivo and in vitro cross-reactivity between Ficus benjamina and Hevea brasiliensis allergens. METHODS: The two patients were asked about use and contact with latex devices and relationship between symptoms and Ficus benjamina exposure. Skin prick tests were performed with Ficus benjamina, Hevea brasiliensis extracts and common allergens. Double-blind nasal and bronchial challenge tests were done using the rinse fluid from a brand of latex gloves. Total and specific IgE antibodies to Ficus benjamina and Hevea brasiliensis were determined. In vitro cross-reactivity was investigated by means of CAP RAST and immunodot inhibition experiments. RESULTS: We observed that for the first patient the primary phenomenon is probably allergy to latex followed by allergy to Ficus benjamina. For the second patient, allergy to Ficus benjamina was diagnosed (improvement related to the avoidance of exposure to Ficus benjamina allergens) and positivity to latex skin prick tests may be due to the cross-reacting allergens. In vitro assays showed specific IgE antibodies to both allergens and cross-reactivity was confirmed in the two cases by reciprocal inhibition of the two extracts. CONCLUSION: The increasing risk of sensitization to widely used latex devices and extensive exposure to Ficus species in households and offices indicates increased allergenic risk from this newly recognized cross-reactivity.
We previously identified a 46-kD protein allergen in latex as having amino acid sequence homology to the patatin gene family. The objective of this study was to characterize this protein by molecular techniques. RNA was isolated from the latex or leaf material from Hevea brasiliensis and from potato tubers. Specific polymerase chain reaction (PCR) primers were designed from the amino acid sequence and reverse transcriptase (RT)-PCR amplified a specific product from latex RNA that was subsequently cloned and sequenced. This product was 1493 bp in length with an 1167 bp open reading frame. The deduced amino acid sequence encodes for a 389 aa protein, pI 4.82 with 43% homology to tobacco patatin. Northern analysis of potato, Hevea leaf, and latex RNA demonstrated the message to be most abundant in latex, weakly present in Hevea leaf, but no hybridization occurred with potato RNA. Patatin has lipid acyl-transferase and PLA2-like activity, suggesting it plays a role as a defence-related protein. Other defence-related proteins in latex such as hevein, glucanase, and hevamine are also allergens. Increased production of defence-related proteins as a result of increased tapping of the rubber trees to meet the demand for latex may explain the increased allergenicity of latex.
While less than 1% of the general population is sensitized to latex, the U.S. Occupational Safety and Health Administration estimates that 8-12% of health-care workers are sensitized. The major source of workplace exposure is powdered natural rubber latex (NRL) gloves. NRL is harvested from HEVEA: brasiliensis trees and ammoniated to prevent coagulation resulting in the hydrolysis of the latex proteins. Prior to use in manufacturing, the latex is formulated by the addition of multiple chemicals. Thus, human exposure is to a mixture of residual chemicals and hydrolyzed latex peptides. Clinical manifestations include irritant contact dermatitis, allergic contact dermatitis (type IV), and type I immediate hypersensitivity response. Type I (IgE-mediated) NRL allergy includes contact urticaria, systemic urticaria, angioedema, rhinitis, conjunctivitis, bronchospasm, and anaphylaxis. Taking an accurate history, including questions on atopic status, food allergy, and possible reactions to latex devices makes diagnosis of type-I latex allergy possible. To confirm a diagnosis, either in vivo skin prick testing (SPT) or in vitro assays for latex-specific IgE are performed. While the SPT is regarded as a primary confirmatory test for IgE-mediated disease, the absence of a U.S. Food and Drug Administration-licensed HEVEA: brasiliensis latex extract has restricted its use in diagnosis. Serological tests have, therefore, become critically important as alternative diagnostic tests. Three manufacturers currently have FDA clearance for in vitro tests, to detect NRL-specific IgE. The commercially available assays may disagree on the antibody status of an individual serum, which may be due to the assay's detecting anti-NRL IgEs to different allergenic NRL proteins. Sensitized individuals produce specific IgE antibody to at least 10 potent HEVEA: allergens, Hev b 1-Hev b 10, each of which differs in its structure, size, and net charge. The relative content and ratios of Hevs in the final allergen preparation most probably could effect diagnostic accuracy. The Hev proteins have been cloned and expressed as recombinant proteins. Sequencing demonstrates both unique epitopes and sequences commonly found in other plant proteins. Sequence homology helps to explain the cross reactivity to a variety of foods experienced by latex allergic individuals. The development of recombinant allergens provides reagents that should improve the diagnostic accuracy of tests for latex allergy. Although clinical and exposure data have been gathered on the factors affecting response in latex-allergic individuals, less is known regarding the development of sensitization. Coupled with in vitro dermal penetration studies, murine models have been established to investigate the route of exposure in the development of latex sensitization. Time-course and dose-response studies have shown subcutaneous, intratracheal, or topical administrations of non-ammoniated latex proteins to induce IgE production. Both in vitro penetration and in vivo studies highlight the importance of skin condition in the development of latex allergy, with enhanced penetration and earlier onset of IgE production seen with experimentally abraded skin. The diagnosis of latex allergy is complicated by these variables, which in turn hinder the development of intervention strategies. Further epidemiological assessment is needed to more explicitly define the scope, trends, and demographics of latex allergy. Diagnostic accuracy can be improved through greater knowledge of proteins involved in the development of latex allergy, and better documentation of the presently available diagnostic tests. In vivo and in vitro models can elucidate mechanisms of sensitization and provide an understanding of the role of the exposure route in latex allergy-associated diseases. Together, these efforts can lead to intervention strategies for reducing latex allergy in the workplace.
30 patients who had experienced contact urticaria after exposure to gloves, contraceptives, balloons or dental cofferdams went through a detailed examination including interviews on: exposure to rubber; local and distant symptoms; provocation tests; skin prick tests with (among other substances) latex extracts from the rubber tree Hevea brasiliensis; patch testing, IgE and RAST analyses. 23 patients (77%) reported additional symptoms from distant organs. 13 belonged to medical occupations; 25 were classified as atopics based on history; 21 had raised IgE-values; 25 were RAST-positive to the extracts from Hevea brasiliensis. 13 had a pre-existing hand dermatitis, of whom 6 were patch test positive to standard allergens. Skin prick tests with extracts from Hevea brasiliensis were the most sensitive diagnostic tool, while the RAST analysis was positive in 25/30 cases. These tests and analyses including RAST were negative in 40 control subjects. Provocation tests are of great value, but difficult to standardize. Several patients reported severe reactions at medical and dental examination/treatment situations (surgical gloves, dental cofferdams), and the risk of life-threatening anaphylactic reactions is emphasized.
Rubber pest mites, Calacarus heveae and Tenuipalpus heveae, reach economic damage levels at the end of the rainy season and the beginning of the dry season in Brazil. Therefore, low humidity adaptation might be an important characteristic for predatory mites to successfully control pest organisms. This study determined the effect of the relative humidity (RH) levels of 30-100% on the hatching of larvae of Amblyseius acalyphus, Euseius citrifolius, Iphiseiodes zuluagai, Metaseiulus camelliae, Agistemus floridanus and Zetzellia malvinae at 25 +/- 0.5 degrees C. These predatory mites are common on rubber trees in the state of São Paulo and might be used for introduction in the major rubber tree production regions in the state of Mato Grosso. At 70% RH or higher, viability was 70% or higher for all species, indicating that their performance might be higher during the rainy season than during the dry season. Eggs of E. citrifolius and M. camelliae presented higher viability at the lower relative humidity levels than those of other species, indicating that these species might have higher chance to persist in the dry season. It is suggested that M. camelliae should be further evaluated for introduction in the state of Mato Grosso, considering that this mite is not yet present in that area.
The objective of this study was to evaluate the oviposition rate and the survivorship of some of the most common predators on plants of the natural vegetation of southeastern Brazil, when confined with different food sources. Forty-four laboratory tests were carried out, each corresponding to a different combination of predator species and food source. The predators tested were: Phytoseiidae--Amblyseius acalyphus, Euseius citrifolius, Iphiseiodes zuluagai, Metaseiulus (Metaseiulus) camelliae, Typhlodromips cananeiensis; Stigmaeidae--Agistemus floridanus, Zetzellia malvinae; Tydeidae--Pronematus sp. For most predators, the highest oviposition rate and survivorship were obtained when the prey was one of the tenuipalpid mites, Brevipalpus phoenicis or Tenuipalpus heveae. With the exception of A. floridanus, the predators had low oviposition rate and survivorship on Calacarus heveae relative to their respective rate on other food sources. Predators had also generally low oviposition rate and survivorship when feeding on Tetranychus urticae, Oligonychus gossypii or Frankliniella sp.
Thanatephorus cucumeris is a ubiquitous fungus responsible for many types of plant diseases worldwide. All isolates from infected Hevea brasiliensis trees secreted pectolytic enzymes; polygalacturonase (PG), pectin lyase (PL) and cellulolytic enzymes; beta-glucosidase and cellobiase in culture. The extracts of the rubber tree leaf tissues, inoculated with T. cucumeris did not show any PG activity. However, PL activity was detected in tissue with the establishment of the infection. The levels of beta-glucosidase, an inherent enzyme in Hevea spp. increased rapidly following infection. However, cellobiase was detected only with the initiation of infection. Molecular weights of PG in all isolates were similar and in the range of 53,000 to 58,000. PL also followed the same pattern showing a molecular weight around 39,000.
BACKGROUND: Allergy to natural rubber latex (NRL) results from exposure to proteins derived from Hevea brasiliensis. Type I latex hypersensitivity is observed in certain occupational and other high-risk groups with frequent exposure to NRL products. This includes health care workers (HCWs), workers in the latex industry, children with spina bifida, and atopic individuals. OBJECTIVES: Early reliable diagnosis and avoidance are required for better patient care. Standardized reagents are not presently available for in vitro and in vivo testing and treatment of patients with latex allergy. However, a number of allergens have been isolated and characterized from Hevea latex and NRL products. Currently, a total of 11 major and minor allergens are designated by the International Allergen Nomenclature Committee. This article reviews the structural and functional characteristics of these latex allergenic proteins. RESULTS: NRL-allergenic proteins include those involved in the biosynthesis of polyisoprene and coagulation of latex rubber elongation factor, small rubber particle protein, prohevein, and patatin. Pathogenesis-related proteins include beta-1,3-glucanases, chitinases, and hevamine; and the structural proteins include microhelix protein complex, proline-rich protein, profilins, enolases, and manganese superoxide dismutase. Recombinant allergens demonstrated skin test reactivity in patients with latex allergy. The minimal level of skin test reactivity was about 70 pg/mL for NRL and 1 ng/mL for recombinant allergens. The use of selected recombinant latex allergens (Hev b 5, Hev b 6, and Hev b 7) in skin prick tests identified 93% of allergic individuals, mainly health care workers. CONCLUSIONS: Recombinant latex allergens are clinically reactive and can be produced in a standardized manner, which could potentially provide safe and sensitive reagents for the diagnosis and treatment of type I latex allergy.
Natural rubber (cis-1,4-polyisoprene) is an isoprenoid compound produced exclusively in plants by the action of rubber transferase. Despite a keen interest in revealing the mechanisms of rubber chain elongation and chain length determination, the molecular nature of rubber transferase has not yet been identified. A recent report has revealed that a 24 kDa protein tightly associated with the small rubber particles of Hevea brasiliensis, therefore designated small rubber particle protein (SRPP), plays a positive role in rubber biosynthesis. Since guayule (Parthenium argentatum Gray) produces natural rubber similar in size to H. brasiliensis, it is of critical interest to investigate whether guayule contains a similar protein to the SRPP. A cDNA clone has been isolated in guayule that shares a sequence homology with the SRPP, thus designated guayule homologue of SRPP (GHS), and the catalytic function of the protein was characterized. Sequence analysis revealed that the GHS is highly homologous in several conserved regions to the SRPP (50% identity). In vitro functional analysis of the recombinant protein overexpressed in E. coli revealed that the GHS plays a positive role in isopentenyl diphosphate incorporation into high molecular weight rubbers as SRPP does. These results indicate that guayule and Hevea rubber trees contain a protein that is similar in its amino acid sequence and plays a role in isopentenyl diphosphate incorporation in vitro, implying that it contributes to the enhancement of rubber biosynthetic activity in rubber trees.
Two clones with homology to the alpha-hydroxynitrile lyase (HNL) cDNA clone, MeHNL10, were isolatedfrom a lambdaEMBL3 cassava (Manihot esculenta Crantz) genomic library. Analysis of the sequences showed that both genomic clones contain HNL genes (MeHNL4, MeHNL24) which are interrupted by two introns. RT-PCR analysis of MeHNL4 shows that it is expressed at high levels in seedling roots and at lower levels in cotyledons and young leaves. The deduced amino acid sequences of MeHNL4, MeHNL10, and MeHNL24 show high sequence identity and homology to the HNL from Hevea brasiliensis whose tertiary structure has been solved at 1.9-A resolution by X-ray crystallography. This high homology allowed the construction of model structures for all of the cassava proteins using the MODELLER program. Homology modeling indicates that the short variable exon 2 encodes the "cap" region which is thought to influence the substrate specificity of the protein. Two hybrid proteins were modeled using the core alpha/beta domain of MeHNL10 and the cap region of either the Hevea HNL or a structurally related Zea protein of unknown function. This analysis suggests that changes in the active site can be engineered by swapping exons.
Palmivorein, a new member of the elicitin family, was purified from the culture filtrate of Phytophthora palmivora isolated from the rubber tree, Hevea brasiliensis. The elicitin was obtained by ammonium sulfate precipitation and further purified using ion-exchange and gel filtration. The molecular weight, isoelectric point, amino acid composition and N-terminal sequences of this molecule are reported and compared to other known elicitins. Palmivorein, as determined by SDS-PAGE, is a small protein of M(r) ca. 10,000. It is classified as an alpha-elicitin according to its acidic pI and the valine residue at position 13. Like other elicitins, the P. palmivora elicitin causes tissue necrosis on tested tobacco leaves. It also causes severe wilting and necrosis of Hevea tissue, and leaves of the susceptible rubber clone (with respect to P. palmivora) are much more sensitive to this elicitin than those that are resistant.
BACKGROUND: No characterized diagnostic natural rubber latex skin testing material is licensed for use in the United States. OBJECTIVE: We have conducted a multicenter clinical skin testing study to document the safety and diagnostic sensitivity and specificity of a candidate Hevea brasiliensis nonammoniated latex (NAL) extract. These data are intended to support the licensing of this reagent for the diagnosis of latex allergy in high-risk populations. METHODS: Three hundred twenty-four subjects (304 adults and 20 children) were classified by their clinical history as having latex allergy (LA group, 124 adults and 10 children) or having no latex allergy (NLA group, 180 adults and 10 children). All subjects provided blood samples and then received sequential puncture skin tests (PSTs) at 1, 100, or 1000 microg/mL protein with a bifurcated needle and NAL (Greer Laboratories) from Malaysian Hevea brasiliensis (clone 600) sap. A 2-stage glove provocation test was used to clarify latex allergy status of individuals with positive history/negative PST result and negative history/positive PST result mismatches. RESULTS: Twenty-four subjects (15%) originally designated as having LA on the basis of their initial clinical history were reclassified to the NLA group on the basis of a negative glove provocation test result. Of the 134 subjects with LA, 54 (40%) were highly sensitive to latex, with a positive PST result at 1 microg/mL NAL. The Greer NAL reagent produced a positive PST rate (sensitivity) of 95% and 99% in subjects with LA at 100 microg/mL and 1 mg/mL, respectively. The negative PST rate (specificity) in 190 subjects with a negative history with the NAL extract at 100 microg/mL and 1 mg/mL, was 100% and 96%, respectively. Immediately after the PST, mild systemic reactions (mainly pruritus) were recorded in 16.1 % of the adults in the LA group and 4.4% of the adults in the NLA group. No reactions required treatment with epinephrine. Only mild delayed reactions were observed in 9.6% (LA group) and 2.8% (NLA group) of subjects 24 to 48 hours after PST. Mean wheal and erythema diameters measured in the 10 children in the LA group with spina bifida at 100 microg/mL and 1 mg/mL were similar to those observed in the adults in the LA group, suggesting that children are not at increased risk for systemic reactions compared with adults. CONCLUSIONS: A suggestive clinical history is necessary but not sufficient for a definitive diagnosis of IgE-dependent latex allergy. These data support the safety and diagnostic efficacy of the Greer NAL, skin test reagent at 100 micro/mL and 1 mg/mL for confirmatory PSTs.
A structural study of low molecular weight rubbers from Jackfruit (Artocarpus heterophyllus) and Painted spurge (Euphorbia heterophylla) was carried out as model compounds of natural rubber from Hevea brasiliensis. The rubber content of latex from Jackfruit was 0.4-0.7%, which is very low compared with that of 30-35% in the latex from Hevea tree. The rubber from Jackfruit latex was low molecular weight with narrow unimodal molecular weight distribution (MWD), whereas that obtained from E. heterophylla showed very broad MWD. The 1H and 13C NMR analyses showed that Jackfruit rubber consists of a dimethylallyl group and two trans-isoprene units connected to a long sequence of cis-isoprene units. The alpha-terminal group of Jackfruit rubber was presumed to be composed of a phosphate group based on the presence of 1H NMR signal at 4.08 ppm corresponding to the terminal =CH-CH2OP group.
To date, 26 species of Phytophthora have been reported in mainland China but only 23 are accepted as good taxa. Phytophthora nicotianae (P. parasitica) is the single most important species causing over 40 different plant diseases; P. boehmeriae is widespread attacking Gossypium, Boehmeria, Citrus, Broussonetia papyrifera and Pterocarya stenoptera; P. "fragariae var. oryzo-bladis" is unique to mainland China causing blight of Oryza sativa seedlings; P. megasperma sensu lato has been isolated from Glycine max, Asparagus officinalis and Spinacia oleracea. In Hainan Province, P. heveae is present in the soil without causing apparent diseases to Hevea brasiliensis there.