Induced resistance in nectarine fruit by Bacillus licheniformis W10 for the control of brown rot caused by Monilinia fructicola.

Brown rot caused by Monilinia fructicola has led to considerable preharvest and postharvest losses in all major nectarine fruit-growing areas. In our previous study, we successfully identified a biocontrol strain of bacteria, Bacillus licheniformis W10, that can be used to control brown rot. However, the possible mechanism of the control of brown rot by B. licheniformis W10 is still unclear. Therefore, the objectives of this study were to determine whether B. licheniformis W10 induces resistance by activating defense-related enzymes including antioxidant enzymes in nectarine. Treatment of nectarine fruit with B. licheniformis W10 reduced both M. fructicola-induced oxidative damage and reactive oxygen species (ROS) production. Furthermore, application of B. licheniformis to nectarine fruit resulted in a significant increase in the activity of antioxidant and defense-related enzymes and increase in the expression of the corresponding genes. Overall, our results verified the proposed mechanism of B. licheniformis W10 in controlling M. fructicola via regulation of ROS levels and activation of antioxidant and defense-related enzymes.

Silencing of one copy of the translation initiation factor eIFiso4G in Japanese plum (Prunus salicina) impacts susceptibility to Plum pox virus (PPV) and small RNA production.

BACKGROUND: In plants, host factors encoded by susceptibility (S) genes are indispensable for viral infection. Resistance is achieved through the impairment or the absence of those susceptibility factors. Many S genes have been cloned from model and crop species and a majority of them are coding for members of the eukaryotic translation initiation complex, mainly eIF4E, eIF4G and their isoforms. The aim of this study was to investigate the role of those translation initiation factors in susceptibility of stone fruit species to sharka, a viral disease due to Plum pox virus (PPV). RESULTS: For this purpose, hairpin-inducing silencing constructs based on Prunus persica orthologs were used to generate Prunus salicina (Japanese plum) 4E and 4G silenced plants by Agrobacterium tumefaciens-mediated transformation and challenged with PPV. While down-regulated eIFiso4E transgenic Japanese plums were not regenerated in our conditions, eIFiso4G11-, but not the eIFiso4G10-, silenced plants displayed durable and stable resistance to PPV. We also investigated the alteration of the si- and mi-RNA profiles in transgenic and wild-type Japanese plums upon PPV infection and confirmed that the newly generated small interfering (si) RNAs, which are derived from the engineered inverted repeat construct, are the major contributor of resistance to sharka. CONCLUSIONS: Our results indicate that S gene function of the translation initiation complex isoform is conserved in Prunus species. We discuss the possibilities of using RNAi silencing or loss-of-function mutations of the different isoforms of proteins involved in this complex to breed for resistance to sharka in fruit trees.

New Data Completing the Spectrum of the Ma, RMia, and RMja Genes for Resistance to Root-Knot Nematodes (Meloidogyne spp.) in Prunus.

Root-knot nematodes (RKN) (Meloidogyne spp.) are worldwide pests that affect a considerable number of plants, among which stone fruit (Prunus spp.) are severely attacked. Prevalent RKN species are Meloidogyne arenaria, M. incognita, and M. javanica in stone fruit but the emergent M. ethiopica and M. enterolobii are also reported to challenge perennial crops. In Prunus spp., the complete-spectrum resistance (R) gene Ma from plum and the more restricted-spectrum R genes RMia from peach and RMja from almond completely inhibit nematode multiplication and gall formation of the RKN species that they control. This study aimed to update the resistance spectra of these three major genes by evaluating their activity toward one isolate of the yet-untested RKN species mentioned above. To state whether a given gene controls a particular species, the principle of our experiment was to genotype with appropriate markers a number of individuals segregating for this gene and then to phenotype these individuals. A perfect matching of the genotype and the phenotype of individuals indicates that the gene of interest is active against and, thus, controls the corresponding isolate of this RKN species. Segregating materials used were an Ma F1 plum progeny, an RMia F2 peach progeny, and an RMja F2 almond progeny. In addition to previous data, our results establish a clear spectrum for each of the three genes toward isolates from both the three prevalent species and the two emerging species. Ultimately, our results reveal that (i) Ma controls all of them, (ii) RMja controls all species except M. incognita and M. floridensis, and (iii) RMia controls M. arenaria, M. incognita, and M. ethiopica but not M. javanica or M. enterolobii. Our data should have wide implications for RKN resistance management and breeding and for deciphering the molecular mechanisms of the spectrum of RKN R genes.

Do Skin Prick Test and In Vitro Techniques Diagnose Sensitization to Peach Lipid Transfer Protein and Profilin Equally Well in Allergy to Plant Food and Pollen?

OBJECTIVE: To compare the skin prick test (SPT) with in vitro techniques (single and multiplex fluorescence enzyme-immunoassay [FEIA]) for detecting sensitization to profilin and lipid transfer protein (LTP). METHODS: We retrospectively studied 181 patients with pollen and/or plant food allergy and 61 controls. SPT was performed with date palm profilin (Pho d 2) and peach LTP (Pru p 3), and specific IgE (sIgE) to Phl p 12 and Pru p 3 was analyzed using single FEIA and microarray. RESULTS: Fifteen of 201 patients with negative results for LTP in the SPT were sensitized to this allergen in the in vitro tests, and 18 of 41 patients with positive results for LTP in the SPT were not sensitized according to the in vitro tests. Seventeen of 186 patients with negative results for profilin in the SPT were sensitized to Phl p 12 by serum sIgE, and 30 out of 56 patients with positive results for profilin in SPT were not sensitized to Phl p 12 according to the other tests. Moderate agreement was observed between the 3 techniques studied. CONCLUSIONS: SPT is a sensitive technique for detecting sensitization to LTP and profilin. Its results are similar to those of in vitro techniques, especially in patients with negative SPT results for peach LTP and palm tree profilin.

Hypersensitivity to Tomato (Lycopersicon esculentum) in Peach-Allergic Patients: rPrup 3 and rPrup 1 Are Predictive of Symptom Severity.

UNLABELLED: Background: The role of allergens in the severity of tomato allergy symptoms has not yet been studied. OBJECTIVES: To evaluate the relationship between severe allergic reactions to peach and tomato and between tomato allergy symptoms and the pattern of IgE positivity for rPru p 1, rPru p 3, rPru p 4, rBetv 1, rBetv 2, rBetv4, rPhl p 1, and rPhl p 12 in order to identify the role of recombinant allergens in the severity of reactions to tomato. METHODS: We studied peach-allergic patients with clinical reactions to tomato by performing an open food challenge, skin prick test, and determination of serum specific IgE to tomato and to recombinant peach, birch, and grass allergens. Statistical analysis was carried out to evaluate the relationship between the severity of tomato symptoms and IgE positivity to the different allergens and to peach-induced symptoms. RESULTS: We found a significant association between severe reactions to tomato and severe reactions to peach (P = .01 7) and levels of IgE to rPru p3 (P = .029) and between mild tomato allergy symptoms and levels of IgE to rPru p1 (P = .047), anti-rBetv 1 (P = .0414), anti-rBetv 2 (P = .0457), and Phleum pratense (P = .0022). CONCLUSION: We observed a significant relationship between peach and symptoms of tomato allergy. IgE positivity for rPru p3 seems to be a surrogate biochemical marker for severe tomato allergy, whereas the presence of anti-rPru p 1 IgE may be an indicator of mild tomato allergy.

Tomato nsLTP as an "In Vivo" Diagnostic Tool: Sensitization in a Mediterranean Population.

BACKGROUND: Tomato allergies have been extensively studied but component-resolved in vivo diagnosis with purified allergens has yet to be performed. OBJECTIVES: To evaluate the prevalence of sensitization to Sola l 3 in a Mediterranean population, and to compare the resulting sensitization profile with that of individuals sensitized to tomato, peach, and/or purified lipid transfer protein (LTP). METHODS: Sola l 3 was purified, characterized, and used to prepare skin prick tests (SPTs). Two groups of patients were selected. Group 1 consisted of patients with at least 1 positive SPT to tomato, peach, or LTP mixture (marker extracts) who were subsequently tested with Sola l 3 (n = 280). Group 2 (prevalence study) consisted of patients who underwent simultaneous SPT with the 3 marker extracts and Sola l 3 (n = 658). Patients from either group who were positive to any of the 4 extracts were studied in detail (study group, n = 1 23). ELISA and immunoblot assays were performed in individuals with a positive SPT to Sola l 3 to detect the presence of specific IgE antibodies to this allergen. RESULTS: Prevalence of sensitization to Sola l 3 was 3.2% overall and 54.7% in tomato-positive patients. Most tomato-sensitized patients were asymptomatic. Symptoms were more common in Sola l 3-positive individuals. Sensitization to peach and the LTP mixture did not discriminate between Sola l 3-positive and Sola l 3-negative patients. CONCLUSIONS: This study confirms that LTP, not only from peach but also from other fruit and vegetables, including tomato, is an important allergen in the Mediterranean area. Sensitization to Sola l 3 is associated with more symptoms in tomato-sensitized patients.

Impact of glutathione on the allergenicity of the peach lipid transfer protein Pru p 3.

BACKGROUND AND OBJECTIVE: The allergenic potential of proteins can be altered under various physicochemical conditions. Glutathione (GSH) is a reducing agent that is used as an antioxidant in food products. We aimed to characterize the natural folding of peach proteins and test the allergenicity of reduced and natural Pru p 3, the major peach allergen. METHODS: Pru p 3 was purified from peach, and its conformation was analyzed by means of circular dichroism. Using a thiol fluorescent probe, reduced proteins were detected in fresh peach. GSH-reduced Pru p 3 was tested in vitro for T-cell proliferation and in vivo using skin prick testing. RESULTS: GSH-reduced Pru p 3 produced variable skin prick reactions in peach-allergic patients. The proliferative response of peripheral blood mononuclear cells from allergic patients to reduced Pru p 3 tended to be less intense, whereas secretion of the cytokines IFN-γ, IL-5, and IL-10 was comparable. In a pool of sera from peach-allergic patients, reduction hardly impaired IgE-binding. Moreover, the stability of reduced Pru p 3 to gastrointestinal digestion was similar to that of the natural form. CONCLUSIONS: GSH can at least transiently reduce Pru p 3. We found that the effect of reduction on the allergenicity of Pru p 3 varied. Therefore, as an additive, GSH does not seem to eliminate the risk of reactions for peach-allergic patients.

Allergen component specific ige measurement with the Immulite™ 2000 system: diagnostic accuracy and intermethod comparison.

BACKGROUND: The identification of the allergenic molecules, associated to the advances in the field of recombinant allergens, led to the development of a new concept in allergy diagnosis called component-resolved diagnosis. The aim of our study was to evaluate the diagnostic accuracy of different allergen components using the full automatic singleplex quantitative platform Immulite™ 2000. METHODS: One hundred ninety-five allergic outpatients (35 to olive pollen, 35 to birch pollen, 35 to profilin, 35 to house dust mites, 35 to peach, and 20 to shrimp) and 20 negative controls were enrolled for the study. Bet v 1, Bet v 2, Ole e 1, Der p 1, Der p 2, Der f 1, Der f 2, Pru p 3, tropomyosin were tested both with Immulite™ 2000 and ImmunoCAP™ (Thermo Fisher Scientific, Uppsala, Sweden). RESULTS: Sensitivity of allergen-specific Immunoglobulin E (sIgE) to Ole e 1, Bet v 1, Der p 1, Der p 2, Der f 1, Der f 2, Pen m 1, and Pru p 3 with Immulite™ 2000 was 100%, 100%, 77.1%, 94.3%, 71.4%, 94%, 75%, and 97.1%, respectively, and the specificity was 100% for all the allergens. The overall agreement between Immulite™ 2000 and ImmunoCAP™ (Thermo Fisher Scientific) platforms was 98.6% (Cohen's kappa = 0.979; confidence interval [CI] 95%: 0.960-0.997). From moderate to strong, positive linear correlations between the assays (r(2) from 0.322 to 0.860, and Spearman's rho from 0.824 to 0.971) were showed. CONCLUSIONS: A high diagnostic accuracy of the sIgE to allergen components measurement with Immulite™ 2000 and a high agreement with ImmunoCAP™ platforms were shown in this study.

Value of microarray allergen assay in the management of eosinophilic oesophagitis.

BACKGROUND: Eosinophilic oesophagitis (EoE) is a disorder characterised by oesophageal dysfunction and, histologically, by eosinophilic inflammation. Although treatment, which includes dilatations, oral corticosteroids and restrictive diets, is often effective, choosing the foods to be eliminated from the diet is difficult. OBJECTIVE: Component resolved diagnostic by microarray allergen assay may be useful in detecting allergens that might be involved in the inflammatory process. METHODS: We studied 67 patients with EoE, diagnosed clinically and histologically by endoscopic biopsy. CRD analysis with microarray technology was carried out in the 67 EoE patients, 50 patients with pollen allergy without digestive symptoms, and 50 healthy controls. RESULTS: Allergies were not detected by microarray in only seven of the 67 patients with EoE. Controls with pollen allergy showed sensitisation to different groups of pollen proteins without significant differences. In EoE patients with response to some allergens, the predominant allergens were grasses group 1 and, in particular, nCyn d 1 (Cynodon dactylon) or Bermuda grass pollen in 59.5%, followed by lipid transfer proteins (LTP) of peach (19.40%), hazelnut (17.91%) and Artemisia (19.40%). CONCLUSIONS: In patients with EoE, sensitisation to plant foods and pollen is important. The proteins most frequently involved are nCyn d 1 and lipid transfer proteins, hazelnuts and walnuts. After one year of an array-guided exclusion diet and pollen-specific immunotherapy in the case of high levels of response, patients with EoE showed preliminary significant improvements.

PPV susceptibility of commonly used peach rootstock-scion combinations.

Sharka disease is one of the most devastating plant epidemics of Prunus species, caused by plum pox virus (PPV). The viral infection affects the fruits by weight-loss and degradation of quality properties. Breeding of resistant rootstocks and cultivars is one of the most effective disease control methods. PPV determines the peach production all over the world. On the world's fruit production list peach is in the sixth, in the Mediterranean region in the fourth place. In this study new data were shown about PPV susceptibility of commonly used rootstock-scion combinations from Hungary. Reverse transcription PCR (RT-PCR) analysis was conducted on the samples from a commercial orchard; the results were evaluated by chi-square test and binary logistic regression. Four rootstock ('GF677', 'PeMa', 'Cadaman' and almond seedlings) and three scion cultivars (Prunus persicae 'Michelini', 'Babygold 6' and 'Cresthaven') were included in this experiment. The rootstocks did not show any significant differences in regard to the resistance of the virus infection (40-50%), but in case of scions, strong significant relations were observed. In case of the combinations there were results in both directions; tolerant and susceptible combinations were observed as well.

Rootstock-to-scion transfer of transgene-derived small interfering RNAs and their effect on virus resistance in nontransgenic sweet cherry.

Small interfering RNAs (siRNAs) are silencing signals in plants. Virus-resistant transgenic rootstocks developed through siRNA-mediated gene silencing may enhance virus resistance of nontransgenic scions via siRNAs transported from the transgenic rootstocks. However, convincing evidence of rootstock-to-scion movement of siRNAs of exogenous genes in woody plants is still lacking. To determine whether exogenous siRNAs can be transferred, nontransgenic sweet cherry (scions) was grafted on transgenic cherry rootstocks (TRs), which was transformed with an RNA interference (RNAi) vector expressing short hairpin RNAs of the genomic RNA3 of Prunus necrotic ringspot virus (PNRSV-hpRNA). Small RNA sequencing was conducted using bud tissues of TRs and those of grafted (rootstock/scion) trees, locating at about 1.2 m above the graft unions. Comparison of the siRNA profiles revealed that the PNRSV-hpRNA was efficient in producing siRNAs and eliminating PNRSV in the TRs. Furthermore, our study confirmed, for the first time, the long-distance (1.2 m) transfer of PNRSV-hpRNA-derived siRNAs from the transgenic rootstock to the nontransgenic scion in woody plants. Inoculation of nontransgenic scions with PNRSV revealed that the transferred siRNAs enhanced PNRSV resistance of the scions grafted on the TRs. Collectively, these findings provide the foundation for 'using transgenic rootstocks to produce products of nontransgenic scions in fruit trees'.

Basophil response to peanut allergens in Mediterranean peanut-allergic patients.

Ara h 1, Ara h 2, and Ara h 3 are important sensitizers in peanut allergy. Ara h 9 has also been shown to be relevant in the Mediterranean area. We evaluated the basophil response to peanut allergens and Pru p 3 in Mediterranean patients: Group 1, peanut and peach allergy; Group 2, peanut allergy and tolerance to peach; Group 3, peach allergy and tolerance to peanut; Group 4, nonallergic subjects that tolerate both peanut and peach. Compared to controls (Group 4), there was an increased basophil activation with Ara h 2 (P = 0.031) and Pru p 3 (P = 0.009) in Group 1 and with Ara h 1 (P = 0.016), Ara h 2 (P = 0.001), and Ara h 9 (P = 0.016) in Group 2. Importantly, only Ara h 2 showed an increased activation (P = 0.009) in Group 2 compared to Group 3. Ara h 2 is the best discriminating allergen for peanut allergy diagnosis in a Mediterranean population showing two patterns: patients also allergic to peach, responding to Ara h 2 and Pru p 3, and patients allergic only to peanut, responding to Ara h 1, Ara h 2, and Ara h 9.

Pomegranate ( Punica granatum L.) expresses several nsLTP isoforms characterized by different immunoglobulin E-binding properties.

BACKGROUND: Pomegranate allergy is associated with sensitization to non-specific lipid transfer proteins (nsLTPs). Our aim was to identify and characterize the non-specific nsLTPs expressed in pomegranate at the molecular level and to study their allergenic properties in terms of immunoglobulin E (IgE)-binding and cross-reactivity with peach nsLTP (Pru p 3). METHODS: A non-equilibrium two-dimensional (2-D) electrophoretic approach based on acid-urea PAGE and sodium dodecyl sulfate PAGE was set up to separate pomegranate nsLTPs. Their immunoreactivity was tested by immunoblotting carried out with anti-Pru p 3 polyclonal antibodies and sera from pomegranate-allergic patients. For final identification, pomegranate nsLTPs were purified by chromatography and subjected to trypsin digestion and mass spectrometry (MS) analysis. For this purpose, the sequences obtained by cDNA cloning of three pomegranate nsLTPs were integrated in the database that was subsequently searched for MS data interpretation. RESULTS: Four nsLTPs were identified by 2-D immunoblotting. The detected proteins showed different IgE-binding capacity and partial cross-reactivity with Pru p 3. cDNA cloning and MS analyses led to the identification of three nsLTP isoforms with 66-68% amino acid sequence identity named Pun g 1.0101, Pun g 1.0201 and Pun g 1.0301. CONCLUSIONS: By 2-D electrophoresis, we could separate different nsLTP isoforms possessing different IgE-binding properties, which might reflect peculiar allergenic potencies. The contribution of Pru p 3 to prime sensitization is not central as in other plant nsLTPs.

Food-specific serum immunoglobulin E measurements in children presenting with food allergy.

BACKGROUND: In children with food allergy, multiple food-specific serum IgE levels to common food allergens are frequently measured. OBJECTIVE: To compare food-specific serum IgE measurements among common food allergens in children with food allergy to determine the characteristics of the measurements, their ability to discriminate between foods associated and not associated with a presenting clinical reaction, and their change over time. METHODS: A retrospective analysis was conducted of food-specific serum IgE to cow's milk, egg white and yolk, peanuts, almond, and soy, for up to 3 subsequent measurements, in 291 children with food allergy. A food-specific serum IgE level lower than 0.35 kU/L was considered a negative measurement. The correlation of IgE measurements with presenting symptoms was conducted for each food in 172 children. RESULTS: Of 1,312 food-specific serum IgE measurements, 69.8% were positive. The median (interquartile range) IgE level for foods associated with the presenting complaint was 7.3 kU/L (2.7-31) and that for foods not associated with a clinical complaint was 2.2 kU/L (0.38-13). The difference was statistically significant (P = .01) only for cow's milk. Specific IgE levels were highest for peanuts, followed by cow's milk, eggs, soy, and almonds, and trended upward over time. CONCLUSION: In children presenting with clinical symptoms of a reaction to a food allergen, measurements of food-specific serum IgE to other common food allergens are commonly positive. An increase in food-specific serum IgE occurs over time.

Are basophil activation and sulphidoleukotriene determination useful tests for monitoring patients with peach allergy receiving sublingual immunotherapy with a Pru p 3-enriched peach extract?

INTRODUCTION: Treatment of food allergy essentially consists of food avoidance, but immunotherapy with food is emerging as a new therapeutic option. OBJECTIVE: To evaluate clinical improvement and immunological changes in patients with peach allergy following sublingual immunotherapy (SLIT) with a Prup3 quantified peach extract. METHODS: A randomized, double-blind, placebo-controlled clinical trial with peach SLIT was conducted. We assessed clinical efficacy after 6 months of treatment by means of double-blind, placebo-controlled oral challenges with peach and also evaluated immunological changes (basophil activation test [BAT] and determination of sulphidoleukotriene production) following stimulation with peach peel and pulp, rPrup3, rMald 1, and rMal d 4 stimulation. We also measured specific IgE and IgG4 to Pru p3. RESULTS: After 6 months of SLIT (T6), the active group showed a 3-fold improvement in tolerance to Prup3 and a significant increase in IgE to rPrup3 and in sLT production following stimulation with peach peel and rPrup3. There was also a significant increase in BAT results after stimulation with rPrup3 at 1 month of SLIT (T1). Statistically significant between-group differences were only observed for BAT with peach peel and pulp at T1 and T6 and for BAT with rPru p3 at T6. No changes were observed in BAT with rMal d 1 or rMal d 4 or in IgG4 levels to nPrup3. CONCLUSIONS: SLIT with a Pru p 3 quantified peach extract is clinically effective and leads to an increase in basophil activation and sulphidoleukotriene production following stimulation with rPru p3 and peach peel in the first months of treatment.

In patients with LTP syndrome food-specific IgE show a predictable hierarchical order.

BACKGROUND: Lipid transfer protein (LTP) is a widely cross-reacting allergen in plant foods. OBJECTIVE: To assess whether IgE to vegetable foods show predictable trends in LTP allergic patients. METHODS: Clinical allergy to plant foods other than peach was sought in 15 consecutive peach-allergic adults monosensitized to LTP. IgE specific for peach, apple, hazelnut, walnut, peanut, lentil, maize, soybean, tomato, sesame, mustard melon, kiwi, and celery as well as to mugwort pollen was measured. RESULTS: Peach-specific IgE levels exceeded IgE to all other study foods. The higher were peach-specific IgE levels, the higher was the probability that other plant-derived foods scored positive. Mean IgE levels specific for all study foods were strongly correlated to peach specific IgE. Food-specific IgE followed a rather precise hierarchy, both in terms of number of positive in-vitro tests and of IgE levels, with apple at the second place after peach, followed by walnut, hazelnut, peanut, lentil, maize, soybean, tomato, kiwi, sesame, mustard, melon, and celery. Such hierarchy was not necessarily paralleled by clinical allergy as lentil, maize, and soybean scored positive in the majority of patients, but induced allergy in 0, 1, and 0 patients, respectively. IgE levels were not necessarily correlated with the severity of clinical allergy. Little or no IgE reactivity to mugwort pollen was found. CONCLUSIONS: In LTP syndrome, IgE reactivity to foods other than peach is in most cases predictable and follows a regular sequence that probably depends on the degree of homology with Pru p3. The reasons why some foods are tolerated by most patients despite elevated IgE reactivity remains to be elucidated.

Diverse amino acid changes at specific positions in the N-terminal region of the coat protein allow Plum pox virus to adapt to new hosts.

Plum pox virus (PPV)-D and PPV-R are two isolates from strain D of PPV that differ in host specificity. Previous analyses of chimeras originating from PPV-R and PPV-D suggested that the N terminus of the coat protein (CP) includes host-specific pathogenicity determinants. Here, these determinants were mapped precisely by analyzing the infectivity in herbaceous and woody species of chimeras containing a fragment of the 3' region of PPV-D (including the region coding for the CP) in a PPV-R backbone. These chimeras were not infectious in Prunus persica, but systemically infected Nicotiana clevelandii and N. benthamiana when specific amino acids were modified or deleted in a short 30-amino-acid region of the N terminus of the CP. Most of these mutations did not reduce PPV fitness in Prunus spp. although others impaired systemic infection in this host. We propose a model in which the N terminus of the CP, highly relevant for virus systemic movement, is targeted by a host defense mechanism in Nicotiana spp. Mutations in this short region allow PPV to overcome the defense response in this host but can compromise the efficiency of PPV systemic movement in other hosts such as Prunus spp.

Peamaclein--a new peach allergenic protein: similarities, differences and misleading features compared to Pru p 3.

BACKGROUND: Among the peach-derived allergens which are already known, the lipid transfer protein (Pru p 3) seems to be the one to exert severe allergic reactions. OBJECTIVE: To identify and characterize a new peach allergen causing a clinical picture similar to that of Pru p 3. METHODS: Patients were selected on the basis of their severe clinical reactivity and negative results to a panel of peach allergens available on the ISAC103 microarray. Several in-house and commercial preparations were compared. Several methods were used to characterize the newly identified molecule. Specific IgE and inhibition assays were performed using the Allergen micro-Beads Array (ABA) assay. RESULTS: Negative ISAC results to Pru p 3 were confirmed by additional testing in contrast with the positive results obtained by commercial Pru p 3-enriched peach peel extracts. The analyses of one of these preparations led to the identification of Peamaclein, a new allergenic protein. It is a small, basic, cysteine-rich, heat-stable, digestion-resistant protein, homologous to a potato antimicrobial peptide. Peamaclein was able to trigger positive skin test reactions and to bind IgE in the ABA assay. It displays an electrophoretic mobility and chromatographic behaviour similar to that of Pru p 3; therefore, it can be hidden in Pru p 3 preparations. In fact, Pru p 3-enriched peach peel extracts were found to contain both Pru p 3 and Peamaclein by means of comparative in vivo testing, and by biochemical and immunochemical assays. Commercially available anti-Pru p 3 polyclonal antibodies were found to have a double specificity for the two molecules. CONCLUSIONS AND CLINICAL RELEVANCE: A new allergen from peach belonging to a new family of allergenic proteins has been identified and characterized. This knowledge on Peamaclein will improve our understanding on the clinical aspects of the peach allergy and the quality of diagnostic reagents.

Understanding the molecular sensitization for Cypress pollen and peach in the Languedoc-Roussillon area.

BACKGROUND: Cypress allergy is a typical winter pollinosis and the most frequent one in the South of France. Main symptoms are rhinitis, conjunctivitis, and asthma. Peach allergy is common too in Southern Europe. Allergic cross-reactions between cypress and peach have been reported, including an oral allergy syndrome. We wanted to investigate whether a cross-reactive allergen between cypress and peach might be responsible for the observed clinical association. METHODS: We analyzed 127 patients included over a 3-month period, outside the pollen season, and we dosed specific IgE levels, for selected, individual allergens. RESULTS: Patients sensitized to peach were mainly positive for the peach-nonspecific lipid-transfer protein. CONCLUSIONS: Profilins or thaumatins could not explain the observed clinical association between cypress and peach.

Rice allergy demonstrated by double-blind placebo-controlled food challenge in peach-allergic patients is related to lipid transfer protein reactivity.

BACKGROUND: The risk factors for sensitisation to rice and the involved allergens are still partially unknown. In this study we evaluated the clinically relevant aspects of rice allergy in DBPCF-positive patients, the major rice allergens, the severity of peach- and rice-induced symptoms in respect to Pru p 3 sensitisation and the role of anti-rPru p 3 IgE levels as a risk factor for rice allergy. METHODS: In 148 peach-allergic subjects, patients with allergic reactions to rice and rice-positive serum IgE were selected. Symptoms were verified by double-blind placebo-controlled food challenges (DBPCFCs), performed at a maximum dosage of 25 g. Rice allergens, identified by IgE immunoblotting, were characterised by N-terminal amino acid sequencing. The relationship between anti-rPru p 3, 1 and 4 IgE levels and rice symptoms were statistically analysed. RESULTS: Eight out of 10 recruited rice-allergic patients had positive DBPCFCs, while 2 patients were not challenged due to their previously documented severe reactions. All patients with rice-induced symptoms were Pru p 3 positive and presented with higher anti-rPru p 3 levels than the rice-sensitised but tolerant patients. A 9-kDa lipid transfer protein, which was highly homologous to Pru p 3, was identified as the major rice allergen and elicited a positive response in all of the patients. Five patients reacted to a putative 15- to 17-kDa rice allergenic protein, and 3 patients reacted to an [alpha]-amylase/subtilisin inhibitor that was approximately 20 kDa. CONCLUSION: Rarely, allergic reactions to rice can arise in patients with peach allergies who are sensitised to Pru p 3, particularly in patients with high anti-rPru p 3 IgE levels.