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PubMed1978

Activation of the alternative pathway of complement by grain. I. C3PA conversion and quantification of complement consumption by rye.

Ground whole rye and airborne rye dust of comparable size distribution were tested for their ability to activate the complement cascade via the alternative pathway. Precipitin-negative pooled normal human serum was incubated with increasing amounts of the two rye dusts. Electrophoresis of the resultant supernatant fluids demonstrated the conversion of the proactivator of the third component of complement to the gamma-migrating activator of the third component. This activation was completely prevented by pre-treating the serum with the chelator EDTA, while pre-treatment with EGTA allowed suboptimal arc conversion, strongly implying that complement was activated via the alternative pathway. Quantification of the supernanant fluids showed dose-dependent complement consumption as defined by both CH100 immunodiffusion and CH50 tube haemolytic techniques. Airborne rye dust showed a greater quantitative potential than ground whole rye for activating the alternative pathway. These results indicate the possibility of the direct action of airborne organic dusts on the induction of inflammatory sequelae in the lungs of both sensitized and unsensitized individuals.

Air Pollutants↗
PubMed1978

Precipitating factors in respiratory allergic disease in Indonesian children.

Skin test results and IgE antibody levels measured by RAST indicate that hypersensitivity to house dust mite (D. pteronyssinus) is a major feature of asthma and allergic rhinitis in Indonesian children. Total serum IgE levels were higher in the allergic than in control children. 60% (twenty-one out of thirty-five) of the asthmatic children and 56% (five out of nine) of the children with allergic rhinitis had IgE antibodies to the helminth Ascaris lumbricoides compared with none out of four control children. A tendency was found for high IgE antibody levels to D. pteronyssinus to occur in association with low IgE antibody levels to A. lumbricoides and vice versa.

Adolescent↗
PubMed1982

Cross-reactivity between Acacia (wattle) and rye grass pollen allergens. Detection of allergens in Acacia (wattle) pollen.

Immunoglobulin E (IgE), directed against components of Acacia (wattle) pollen, has been detected by radioallergosorbent tests (RAST) in the sera of some children and adults who develop allergic symptoms in the presence of flowering Acacia trees in Australia. All these subjects also had high levels of IgE directed against Lolium perenne (rye grass) pollen. Inhibition by RAST showed that most of the IgE molecules which bound to Acacia pollen components also bound to L. perenne pollen extracts, and to Glycoprotein 1, the major allergen of L. perenne pollen. In these assays, the allergens have been immobilized on polyvinyl chloride microtitre trays: the sensitivity of this approach is compared to that of commercial RAST kits.

Acacia↗
PubMed1982

Predicting the results of allergen bronchial challenge by simple clinical methods.

In this study, the ability to predict allergenic bronchial reactivity from non-specific bronchial reactivity, measured by free corridor-running, and the level of skin reactivity to whole rye grass pollen extract (WRGPE--Lolium perene) and house-dust mite extract (HDM--Dermatophagoides pteronyssinus) was examined in asthmatic children. Discriminant analysis of the data showed that, for the concentration of extracts used and the conditions of bronchial challenge for both allergens, the level of skin sensitivity to undiluted extract and the percentage fall in peak flow-rate on exercise best-predicted the outcome of bronchial challenge for WRGPE (100% of cases) and for HDM (89% of cases). The results suggest that it may be possible to predict allergenic bronchial-reactivity to different allergens in children by simple clinical methods, if conditions of provocation and potency of allergen extracts can be standardized.

Allergens↗
PubMed1982

Immunoglobulin E antibodies to ingested cereal flour components: studies with sera from subjects with asthma and eczema.

The specificity of immunoglobulin E for different cereal grain proteins was investigated using sera from twenty paediatric patients with asthma and/or eczema. Close correlations were observed between radioallergosorbent test values for grain extracts of wheat, rye and barley, and, to a lesser extent, oats. Of the different wheat four fractions tested, the globulins and glutenins consistently bound higher levels of IgE than the gliadins and albumins. This is in contrast both with bakers' asthma (an allergy to inhaled flour where the albumins are important allergens) and with coeliac disease (in which gliadin is the most toxic fraction). Partial digestion of the flour proteins largely removed their ability to bind IgE. An analytical technique of identifying allergens after gel isoelectric focusing demonstrated that many different flour proteins were involved.

Adolescent↗
PubMed1983

Baker's asthma. Clinical and immunological studies.

Seven bakers with respiratory symptoms were evaluated by skin tests, RAST assay for specific IgE antibodies to rye and wheat, inhalation challenge with methacholine for the determination of non-specific bronchial reactivity, and bronchoprovocation with rye and wheat extracts for the determination of antigen-specific bronchial reactivity. An immediate asthmatic response to antigen challenge was observed in four subjects and all of them had a high level of flour-specific IgE antibodies. The serum RAST values provided a more accurate predictive value than the degree of cutaneous sensitivity determined by skin testing with respect to the bronchial response to antigenic challenge. Among those who reacted positively to antigenic bronchoprovocation, a much lower antigen dose was required to elicit a positive reaction if the subject also had an increased degree of non-specific bronchial reactivity. An elevated RAST value was not found in thirty-eight asymptomatic bakers or in ten asthmatics who had no occupational exposure to flour. Thus, baker's asthma appears to be a form of allergic asthma to cereal flours mediated by specific IgE antibodies. Both the level of serum IgE antibodies and the degree of non-specific bronchial reactivity are important factors which may influence a baker's bronchial response upon inhalation of cereal flours.

Adult↗
PubMed1994

Immediate hypersensitivity to bakery, brewery and wine products in yeast-sensitive atopic dermatitis patients.

Ultrafiltered (> 1000 Da) samples of beer, aged red wine, young white wine, sparkling wine and extracts of fresh wheat bread and dried rye bread were analysed by skin-prick test (SPT), radioallergosorbent test (RAST) inhibition, sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting to find out if they contain Saccharomyces cerevisiae (S. cerevisiae, baker's yeast) allergens. Serum pool consisting of S. cerevisiae positive sera was used in the assays. The results were compared with freeze-dried reference S. cerevisiae and cereal antigens. The beer, bread, red wine and sparkling wine extracts elicited immediate reactions. However, no evident correlation with suspected symptoms was observed. White wine extract caused reactions in four out of six atopic dermatitis (AD) patients with symptoms, and in five out of seven symptom-free AD patients and in two of the 24 controls. In SDS-PAGE, protein bands were found in wheat and rye bread extracts and beer. In IgE immunoblotting, however, no staining was seen with the S. cerevisiae positive sera suggesting that they were of cereal origin. In white wine and champagne extracts a non-specific staining was seen in the region 20 kDa representing, e.g. lectin-like activity. No baker's yeast antigen could be detected in brewery and bakery products with IgE-immunoblotting even in the excessively concentrated extracts. The IgE mediated allergy to baker's yeast alone should thus not lead to denial of bakery, brewery and wine products.

Adolescent↗
PubMed1995

Identification of crossreacting wheat, rye, barley and soya flour allergens using sera from individuals with wheat-induced asthma.

We have conducted radio allergosorbent test (RAST), competitive RAST inhibition, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting using sera from patients with wheat-induced asthma to investigate the immunological relationship between wheat, rye, barley and soya, and to identify common proteins between these flours. RAST showed strong associations between the levels of specific IgE to wheat flour and those of rye and barley flour. Competitive RAST inhibition showed that wheat, rye, barley and soya flours contained crossreacting proteins, in decreasing concentrations. Wheat, rye and barley flours had similar protein profiles on gel electrophoresis. Soya flour contained a number of high molecular weight proteins not present in the other cereals. Western blotting using sera from 21 wheat flour hypersensitive individuals identified a large number of allergens in the different flours. Proteins of 69, 33, 26, 21 and 12 kDa were identified as major wheat flour allergens. Rye flour proteins of 21 and 12 kDa, and barley flour proteins of 69, 52 and 10 kDa were the major allergens identified by serum from wheat hypersensitive individuals. The major common protein of soya and wheat flour had a molecular weight of 21 kDa. The majority of crossreacting allergens identified between the different flours have molecular weights similar to those of known flour enzymes or enzyme inhibitors.

Allergens↗
PubMed2006

Airborne exposure to wheat allergens: measurement by human immunoglobulin G4 and rabbit immunoglobulin G immunoassays.

BACKGROUND: Exposure to airborne wheat allergens in the bakery trade is associated with a high risk of occupational allergy and asthma. Control and reduction of allergen exposure require relatively simple but reliable monitoring techniques. We developed new rabbit IgG-based enzyme immunoassays (EIA) for wheat allergens, which might be a convenient alternative for the thus far used human IgG4 inhibition assay. METHODS: The reactivity and specificity of rabbit antibodies were assessed by EIA and immunoblotting, and compared with those of IgE from wheat-sensitized bakers, and with the antibodies used in the IgG4 inhibition EIA. An IgG inhibition and a sandwich EIA were developed for analysis of airborne dust samples. RESULTS: Human IgG4 and rabbit IgG inhibition EIAs had comparable sensitivities, with limits of detection (LOD) between 18 and 88 ng/mL, while the sandwich EIA was much more sensitive (LOD<0.2 ng/mL). Human IgG4 and rabbit IgG reacted in immunoblotting with most of the IgE-binding wheat proteins, although with quantitative differences. All three assays showed a strong reaction with wheat proteins, and some cross-reactivity with rye and barley, but were further highly specific for cereal flour proteins. Concentrations measured with the three EIAs in 432 airborne dust samples were highly correlated (r>0.95) and their absolute values showed less than 10-20% differences. CONCLUSION: The rabbit IgG EIAs are valid substitutes for the human IgG4 inhibition EIA, with important practical advantages. The inhibition EIA is recommended for routine wheat allergen measurements. The sandwich EIA may be used to measure low allergen levels, as in short task-related exposure measurements or in subfractions of airborne dust samples.

Air Pollutants, Occupational↗
PubMed1997

Viability of Cryptosporidium parvum during ensilage of perennial ryegrass.

The survival of Cryptosporidium parvum during ensilage of perennial ryegrass was examined in laboratory silos with herbage prepared in one of three different ways; either untreated, inoculated with a strain of Lactobacillus plantarum or by direct acidification with formic acid. The pH values of all silages initially fell below 4.5, but only formic acid-treated silage remained stable at less than pH 4 after 106 d, with the pH of the untreated and inoculant-treated silages rising to above 6. The formic acid-treated silage had a high lactic acid concentration (109 g kg-1 dry matter (DM)) and low concentrations of propionic and butyric acids after 106 d. However, the untreated and inoculant-treated silages showed an inverse relationship, with low lactic acid concentrations and high concentrations of acetic, propionic and butyric acids. These silages also contained ammonia-N concentrations in excess of 9 g kg-1 DM. In terms of the viability of Cryptosporidium parvum oocysts very few differences were seen after 14 d of ensilage with ca 50% remaining viable, irrespective of treatment and total numbers had declined from the initial level of 5.9 x 10(4) to 1 x 10(4) g(-1) fresh matter. Total oocyst numbers remained approximately the same until the end of the ensiling period, with the percentage of viable oocysts declining to 46, 41 and 32% respectively for formic acid, inoculant and untreated silages. The results are discussed in terms of changes occurring during the silage fermentation, in particular the products which may influence the survival of Cryptosporidium and implications for agricultural practice and the health of silage fed livestock.

Animal Feed↗
PubMed2006

Persistence and efficacy of Beauveria brongniartii strains applied as biocontrol agents against Melolontha melolontha in the Valley of Aosta (northwest Italy).

AIMS: To monitor and select genetically characterized strains of Beauveria brongniartii to be used as microbiological control agents against Melolontha melolontha in different climatic conditions of the Valley of Aosta (northwest Italy). METHODS AND RESULTS: Molecular random amplified polymorphic DNA markers allowed monitoring of five B. brongniartii strains (C2, F, K2, N3 and W2) in field trials. Ten sites were chosen at Jovençan, Saint-Pierre and Quart areas, where a mixture of the five strains colonizing rye kernels was applied to the soil of each M. melolontha infested site. Growth, persistence and virulence on M. melolontha larvae of five fungal strains were evaluated in two subsequent 24-month studies. Beauveria brongniartii grew best at the Jovençan sites. Not only did strain F persist better than the other strains in most soil samples but it was also the most virulent strain. Strain F was isolated the most frequently from infected M. melolontha larvae recovered from the test sites. A general decrease in the larvae rate was detected in the test field soil. CONCLUSIONS: Strain F of B. brongniartii was better than other strains in growth, persistence and virulence against M. melolontha larvae in the test site soil. SIGNIFICANCE AND IMPACT OF THE STUDY: Results obtained from preliminary field trials support the use of strain F as a biological control agent against M. melolontha in the Valley of Aosta even if further targeted studies are still necessary.

Animals↗
PubMed2006

Molecular identification, heterologous expression and properties of light-insensitive plant catalases.

Most catalases are inactivated by light in a heme-sensitized and O2-dependent reaction. In leaves of the alpine plant Homogyne alpina and in the peroxisomal cores of Helianthus annuus, light-insensitive catalases were observed. For the catalases Hacat1 of H. alpina and HnncatA3 of H. annuus, cDNA clones were obtained. Expression of recombinant active enzymes in insect cells confirmed that they coded for light-insensitive catalases. Kinetic and catalytic properties of light-sensitive or light-insensitive catalases did not differ substantially. However, the specific activity of the latter was markedly lower. The light-insensitive catalase HaCAT-1 was not resistant against inactivation by superoxide. Amino acid sequences of the light-insensitive catalases HaCAT-1 and HNNCATA3 were highly identical. They showed only a few exceptional amino acid substitutions at positions that are highly conserved in other catalases. These appeared to be localized mainly in a surface cavity at the entrance of a minor channel leading to the central heme, suggesting that this region played some, though yet undefined, role for light sensitivity. While the replacement of a highly conserved His by Thr225 was the most unique substitution, a single exchange of His225 by Thr in the light-sensitive catalase SaCAT-1 by mutagenesis was not sufficient to reduce its sensitivity to photoinactivation.

Amino Acid Sequence↗
PubMed2004

High-resolution FISH on super-stretched flow-sorted plant chromosomes.

A novel high-resolution fluorescence in situ hybridisation (FISH) strategy, using super-stretched flow-sorted plant chromosomes as targets, is described. The technique that allows longitudinal extension of chromosomes of more than 100 times their original metaphase size is especially attractive for plant species with large chromosomes, whose pachytene chromosomes are generally too long and heterochromatin patterns too complex for FISH analysis. The protocol involves flow cytometric sorting of metaphase chromosomes, mild proteinase-K digestion of air-dried chromosomes on microscopic slides, followed by stretching with ethanol:acetic acid (3 : 1). Stretching ratios were assessed in a number of FISH experiments with super-stretched chromosomes from barley, wheat, rye and chickpea, hybridised with 45S and 5S ribosomal DNAs and the [GAA]n microsatellite, the [TTTAGGG]n telomeric repeat and a bacterial artificial chromosome (BAC) clone as probes. FISH signals on stretched chromosomes were brighter than those on the untreated control, resulting from better accessibility of the stretched chromatin and maximum observed sensitivity of 1 kbp. Spatial resolution of neighbouring loci was improved down to 70 kbp as compared to 5-10 Mbp after FISH on mitotic chromosomes, revealing details of adjacent DNA sequences hitherto not obtained with any other method. Stretched chromosomes are advantageous over extended DNA fibres from interphase nuclei as targets for FISH studies because they still retain chromosomal integrity. Although the method is confined to species for which chromosome flow sorting has been developed, it provides a unique system for controlling stretching degree of mitotic chromosomes and high-resolution bar-code FISH.

Chromosomes, Artificial, Bacterial↗
PubMed1980

Hypersensitivity to inhaled flour allergens. Comparison between cereals.

Radioallergosorbent testing (RAST) of sera from subjects sensitized to wheat and rye flour indicated that there is significant reaction with seed extracts of 12 cereals (wheat, durum wheat, triticale, cereal rye, barley, rye grass, oats, canary grass, rice, maize, sorghum and Johnson grass). Results were evaluated in terms of taxonomic relationships and of the electrophoretically determined protein composition of the cereal extracts. RAST uptakes were uniformly low in sera from four rhinitic bakers, yet were significantly above the levels for non-allergic and cord sera. Much higher RAST uptakes were obtained with sera from four asthmatic bakers when tested with wheat and its close relatives, but there was still reasonably high reactivity with more distantly related cereals. RAST inhibition experiments indicated in a more direct way the extent of cross-reactivity between grain extracts of wheat, rye, barley and oats. One baker had a history of more severe attacks of breathlessness following inhalation of rye flour compared with wheat flour. This was confirmed by bronchial challenge testing, but the comparison was not obviously consistent with the results of prick testing or estimation of histamine released from his leucocytes. The results as a whole suggested that the bran layers of cereal grains are at least as allergenic as flour.

Allergens↗
PubMed1993

Common allergenic structures in hazelnut, rye grain, sesame seeds, kiwi, and poppy seeds.

Allergy to kiwi, poppy seeds, and/or sesame seeds often occurs in patients with a simultaneous sensitization to nuts and flour. Previously cross reactions have been verified by RAST inhibition. In this study the nature of this cross-reactivity is further characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), followed by immunoblotting to nitrocellulose. The degree of cross-reactivity among kiwi, sesame seeds, poppy seeds, hazelnuts, and rye grain was found to be very high in the patients studied. The existence of both cross-reacting and unique components was observed; however, the cross-reacting and unique components could be different for different patients.

Adolescent↗
PubMed1996

Interaction of allergens from house-dust mite and from cereal flours: Dermatophagoides pteronyssinus alpha-amylase (Der p 4) and wheat and rye alpha-amylase inhibitors.

Major wheat and rye allergens associated with baker's asthma have been identified as specific inhibitors of Dermatophagoides pteronyssinus alpha-amylase (Der p 4). the most active inhibitors were the wheat tetrameric one and the rye dimeric RDAI-1. these data suggest that interaction between mite and cereal allergens (alpha-amylase/inhibitor complexes) might occur in house-dust-mite-infected flours. In contrast to the strong inhibition of the d. pteronyssinus enzyme by the wheat and rye inhibitors, the amylases from D. farinae, Lepidoglyphus destructor, and Tyrophagus putrescentiae were much less affected. Moreover, the D. pteronyssinus amylase showed higher pH optimum (6.5 versus 5.0) and activity level (2-6 times/mg of protein in whole-mite extracts) than those from the three other mite species.

Allergens↗
PubMed2004

Rye flour induces a stronger early bronchial response than wheat flour in occupational asthma.

BACKGROUND: Our aims were to compare the doses of wheat and rye flour that induce early bronchial responses in occupationally exposed asthmatic subjects and to assess the effects of the dose of inhaled flour, the duration of exposure and the dose rate. METHODS: Ten patients underwent tests with lactose, wheat flour and rye flour. We compared the decrease in forced expiratory volume in 1 s (FEV(1)) observed during the challenge with flour and with lactose. We also calculated the amount of flour that was instantaneously active. RESULTS: Seven subjects had significantly decreased FEV(1) values following exposure to wheat and rye flour and two subjects only did so for rye flour. The provocative dose (PD, dose required to reduce FEV(1) by 15%) of rye was lower than that of wheat flour (geometric mean; PD(15) rye: 95 microg; wheat: 368 microg). The calculated doses of rye and wheat flour were better correlated with the change in FEV(1) than were the cumulative doses. CONCLUSION: The bronchial response was greater with rye than with wheat flour. The response was related to the dose of allergen inhaled and to the dose rate.

Asthma↗