Immunostimulant plant proteins: Potential candidates as vaccine adjuvants.

The COVID-19 pandemic is shaking up global scientific structures toward addressing antibiotic resistance threats and indicates an urgent need to develop more cost-effective vaccines. Vaccine adjuvants play a crucial role in boosting immunogenicity and improving vaccine efficacy. The toxicity and adversity of most adjuvant formulations are the major human immunization problems, especially in routine pediatric and immunocompromised patients. The present review focused on preclinical studies of immunoadjuvant plant proteins in use with antiparasitic, antifungal, and antiviral vaccines. Moreover, this report outlines the current perspective of immunostimulant plant protein candidates that can be used by researchers in developing new generations of vaccine-adjuvants. Future clinical studies are required to substantiate the plant proteins' safety and applicability as a vaccine adjuvant in pharmaceutical manufacturing.

The Flagellin:Allergen Fusion Protein rFlaA:Betv1 Induces a MyD88- and MAPK-Dependent Activation of Glucose Metabolism in Macrophages.

TLR5 ligand flagellin-containing fusion proteins are potential vaccine candidates for many diseases. A recombinant fusion protein of flagellin A and the major birch pollen allergen Bet v 1 (rFlaA:Betv1) modulates immune responses in vitro and in vivo. We studied the effects of rFlaA:Betv1 on bone marrow-derived macrophages (BMDMs). BMDMs differentiated from BALB/c, C57BL/6, TLR5-/-, or MyD88-/- mice were pre-treated with inhibitors, stimulated with rFlaA:Betv1 or respective controls, and analyzed for activation, cytokine secretion, metabolic state, RNA transcriptome, and modulation of allergen-specific Th2 responses. Stimulation of BMDMs with rFlaA:Betv1 resulted in MyD88-dependent production of IL-1ß, IL-6, TNF-α, IL-10, CD69 upregulation, and a pronounced shift towards glycolysis paralleled by activation of MAPK, NFκB, and mTOR signaling. Inhibition of either mTOR (rapamycin) or SAP/JNK-MAPK signaling (SP600125) resulted in dose-dependent metabolic suppression. In BMDM and T cell co-cultures, rFlaA:Betv1 stimulation suppressed rBet v 1-induced IL-5 and IL-13 secretion while inducing IFN-γ production. mRNA-Seq analyses showed HIF-1a, JAK, STAT, phagosome, NLR, NFκB, TNF, TLR, and chemokine signaling to participate in the interplay of cell activation, glycolysis, and immune response. rFlaA:Betv1 strongly activated BMDMs, resulting in MyD88-, MAPK-, and mTOR-dependent enhancement of glucose metabolism. Our results suggest macrophages are important target cells to consider during restauration of allergen tolerance during AIT.

Reduced anaphylactic potential of denatured pullulan-conjugated Cry j 1.

Japanese Cedar (JC) pollinosis is the most common seasonal allergic rhinitis in Japan. Throughout the JC pollen season, patients suffer from the allergic symptoms, resulting in a reduction of quality of life. Allergy immunotherapy (AIT) is an established treatment option for a wide range of allergens that unlike symptomatic treatments (e.g. antihistamines) may provide sustained immune tolerance. However, AIT, especially subcutaneous immunotherapy (SCIT) has a fatal anaphylaxis risk due to the use of crude allergen extracts. Consequently, development of allergen derivatives with substantially reduced anaphylactic potential is desirable. An allergen derivative that showed reduced IgE-binding and anaphylactic potential was developed through conjugation of native Cry j 1 (n Cry j 1), a major JC allergen, to the polysaccharide pullulan followed by chemical but non-covalent denaturation. The resulting Cry j 1 allergen derivative, Dn p-Cry j 1, showed reduced IgE-binding and IgE-mediated effector cell activation in vitro using an ELISA competition assay and a mast cell activation model (EXiLE). Reduced anaphylactic potential of Dn p-Cry j 1 in vivo was demonstrated using the rat passive cutaneous anaphylaxis (PCA) assay. The difference in anaphylactic potential of Dn p-Cry j 1 compared to n Cry j 1 in wild-type rats was of the same magnitude as the difference seen in the anaphylaxis reactions obtained with n Cry j 1 in wild-type rats and mast-cell deficient rats, indicating a dramatic reduction in anaphylactic potential of Dn p-Cry j 1. These results indicate that Dn p-Cry j 1 is a promising candidate for next-generation JC AIT.

Activation of defense response in common bean against stem rot disease triggered by Trichoderma erinaceum and Trichoderma viride.

White mold and stem rot is a common disease of Phaseolus vulgaris caused by Sclerotinia sclerotiorum. Biological control is a promising alternative for the control of this disease. In the present study, two Trichoderma spp., T. erinaceum and T. viride, and the consortium of both were evaluated as biocontrol agents against sclerotinia stem rot disease. The results revealed that T. erinaceum (NAIMCC-F-02171) and T. viride (NAIMCC-F-02500) when applied alone, significantly suppressed the infection rate of S. sclerotiorum and increased the rate of survival of plants by 74.5%. On the contrary, the combination of both the Trichoderma spp. was found to be more effective in reducing stem rot by 57.2% and increasing the survival of plants by 87.5% when compared to the individual Trichoderma applications. Further, the exogenous supplementation of Trichoderma activated antioxidative machineries, such as peroxidase, polyphenol oxidase, superoxide dismutase, catalase, and ascorbic acid in the plant. Besides, hydrogen peroxide and superoxide-free radical accumulation were also found to be reduced when T. erinaceum and T. viride were used either individually or in combination under the pathogen-challenged condition. Additionally, the photopigments in the bioprimed plants were markedly increased. Moreover, the combined inoculation of the two isolates yielded the highest records of growth parameters (root weight, shoot length, and leaf weight) compared with individual inoculation. Therefore, based on the above results, it was concluded that the combination of T. erinaceum and T. viride can be effectively used as an alternative to control white mold and stem rot caused by S. sclerotiorum.

JA signal-mediated immunity of Dendrobium catenatum to necrotrophic Southern Blight pathogen.

BACKGROUND: Dendrobium catenatum belongs to the Orchidaceae, and is a precious Chinese herbal medicine. In the past 20 years, D. catenatum industry has developed from an endangered medicinal plant to multi-billion dollar grade industry. The necrotrophic pathogen Sclerotium delphinii has a devastating effection on over 500 plant species, especially resulting in widespread infection and severe yield loss in the process of large-scale cultivation of D. catenatum. It has been widely reported that Jasmonate (JA) is involved in plant immunity to pathogens, but the mechanisms of JA-induced plant resistance to S. delphinii are unclear. RESULTS: In the present study, the role of JA in enhancing D. catenatum resistance to S. delphinii was investigated. We identified 2 COI1, 13 JAZ, and 12 MYC proteins in D. catenatum genome. Subsequently, systematic analyses containing phylogenetic relationship, gene structure, protein domain, and motif architecture of core JA pathway proteins were conducted in D. catenatum and the newly characterized homologs from its closely related orchid species Phalaenopsis equestris and Apostasia shenzhenica, along with the well-investigated homologs from Arabidopsis thaliana and Oryza sativa. Public RNA-seq data were investigated to analyze the expression patterns of D. catenatum core JA pathway genes in various tissues and organs. Transcriptome analysis of MeJA and S. delphinii treatment showed exogenous MeJA changed most of the expression of the above genes, and several key members, including DcJAZ1/2/5 and DcMYC2b, are involved in enhancing defense ability to S. delphinii in D. catenatum. CONCLUSIONS: The findings indicate exogenous MeJA treatment affects the expression level of DcJAZ1/2/5 and DcMYC2b, thereby enhancing D. catenatum resistance to S. delphinii. This research would be helpful for future functional identification of core JA pathway genes involved in breeding for disease resistance in D. catenatum.

Design of peptides with high affinity binding to a monoclonal antibody as a basis for immunotherapy.

About half of the US population is sensitized to one or more allergens, as found by a National Health and Nutrition Examination Survey (NHANES). The most common treatment for seasonal allergic responses is the daily use of oral antihistamines, which can control some of the symptoms, but are not effective for nasal congestion, and can be debilitating in many patients. Peptide immunotherapy is a promising new approach to treat allergic airway diseases. The small size of the immunogens cannot lead to an unwanted allergic reaction in sensitized patients, and the production of peptides with sufficient amounts for immunotherapy is time- and cost-effective. However, it is not known what peptides are the most effective for an immunotherapy of allergens. We previously produced a unique monoclonal antibody (mAb) E58, which can inhibit the binding of multiple groups of mAbs and human IgEs from patients affected by the major group 1 allergens of ragweed (Amb a 1) and conifer pollens (Jun a 1, Cup s 1, and Cry j 1). Here, we demonstrated that a combined approach, starting from two linear E58 epitopes of the tree pollen allergen Jun a 1 and the ragweed pollen allergen Amb a 1, and residue modifications suggested by molecular docking calculations and peptide design could identify a large number of high affinity binding peptides. We propose that this combined experimental and computational approach by structural analysis of linear IgE epitopes and peptide design, can lead to potential new candidates for peptide immunotherapy.

Allergy to Gibberellin-Regulated Proteins (Peamaclein) in Children.

BACKGROUND: Gibberellin-regulated proteins (GRPs, Peamaclein) are allergens recently identified in plant-derived food allergy (FA), and little is known about the clinical manifestations of this allergic condition in the European population, especially in children. OBJECTIVE: Our study aimed to identify and characterize pediatric patients with pollen-FA due to GRP sensitization. METHODS: We retrospectively analyzed the charts of patients referred to the Allergy Unit of the Meyer Children's Hospital in Florence for suspected FA. Three main eligibility criteria based on the actual knowledge of GRP allergy were used to select patients deserving further investigations: (1) systemic reactions after consumption of fruit or an unknown culprit food, (2) positive skin prick tests to both cypress pollen and Pru p 3-enriched peach peel extracts, (3) negative in vitro test results for Pru p 3 serum-specific Immunoglobulin E (sIgE). We performed the in vitro test to determine the anti-rPru p 7 (Peamaclein) sIgE levels in the selected patients. RESULTS: We identified 10 pediatric patients with Pru p 7 allergy and described their characteristics. The use of our eligibility criteria showed a high accuracy in identifying these patients: 100% of the selected patients had positive in vitro results for Pru p 7. We therefore proposed a diagnostic algorithm for Pru p 7 allergy. CONCLUSION: This is the first case series of European pediatric patients with a demonstrated Peamaclein allergy. These findings broaden our knowledge on GRP allergy in pediatric populations and could help clinicians to suspect, diagnose, and manage this recently discovered plant-derived FA.

Identification of a vicilin-like major allergen from Prosopis juliflora exhibiting cross- reactivity with legume food allergens.

BACKGROUND: Prosopis juliflora is a clinically relevant allergic sensitizer worldwide and shares cross-reactivity with allergens from several tree pollen and food. The present study aims to purify and immunobiochemically characterize a major allergen from Prosopis pollen. The allergen was further investigated for its cross-reactivity with legume allergens. METHODS: Prosopis extract was fractionated by Q Sepharose and Superdex 75 gel filtration column to purify the allergen. Specific IgE against purified protein was estimated via ELISA and immunoblot. The protein was subjected to mass spectrometric analysis. Glycan characterization was performed by Schiff staining and lectin binding assay followed by deglycosylation studies. The functional activity of the purified protein was evaluated by the basophil activation test. Cross-reactivity was assessed by inhibition studies with legume extracts. RESULTS: A 35 kDa protein was purified and showed 75% IgE reactivity with the patients' sera by ELISA and immunoblot. Glycan characterization of protein demonstrated the presence of terminal glucose and mannose residues. A reduction of 40% and 27% in IgE binding was observed upon chemical and enzymatic deglycosylation of the protein, respectively. The glycoprotein allergen upregulates the expression of CD203c on basophils which was significantly reduced upon deglycosylation, signifying its biological ability to activate the effector cells. The identified protein shared significant homology with Lup an 1 from the lupine bean. Immunoblot inhibition studies of the purified allergen with legume extracts underlined high cross-reactive potential. Complete inhibition was observed with peanut and common bean, while up to 70% inhibition was demonstrated with soy, black gram, chickpea, and lima bean. CONCLUSION: A 35 kDa vicilin-like major allergen was isolated from P. juliflora. The protein possesses glycan moieties crucial for IgE binding and basophil activation. Furthermore, the purified protein shows homology with Lup an 1 and exhibits cross-reactivity with common edible legume proteins.

Biochemical composition of Phleum pratense pollen grains: A review.

The Poaceae family is composed of 12,000 plant species. Some of these species produce highly allergenic anemophilous pollen grains (PGs). Phleum pratense pollen grains (PPPGs) emerged as a model for studies related to grass allergy. The biochemical composition of allergenic PGs has not yet been fully described despite potential health effects of PG constituents other than allergenic proteins. This review brings together the information available in literature aiming at creating a comprehensive picture of the current knowledge about the chemical composition of allergenic PGs from timothy grass. PPPGs have an average diameter between 30-35 µm and the mass of a single PG was reported between 11 and 26 ng. The pollen cytoplasm is filled with two types of pollen cytoplasmic granules (PCGs): the starch granules and the polysaccharide particles (p-particles). Starch granules have a size between 0.6-2.5 µm with an average diameter of 1.1 µm (estimated number of 1000 granules per PG) while p-particles have a size ranging around 0.3 to 0.4 µm (estimated number between 61,000-230,000 p-particles per PG). The rupture of PG induces the release of PCGs and the dispersion of allergens in the inhalable fraction of atmospheric aerosol. PPPGs are composed of sporopollenin, sugars, polysaccharides, starch, glycoproteins (including allergens), amino-acids, lipids, flavonoids (including isorhamnetin), various elements (the more abundant being Si, Mg and Ca), phenolic compounds, phytoprostanoids, carotenoids (pigments) metals and adsorbed pollutants. PPPG contains about a hundred different proteins with molecular masses ranging from 10 to 94 kDa, with isoelectric points from 3.5-10.6. Among these proteins, allergens are classified in eleven groups from 1 to 13 with allergens from groups 1 and 5 being the major contributors to Phl p pollen allergy. Major allergen Phl p 5 was quantified in PPPGs by several studies with concentration ranging from 2.7 and 3.5 µg.mg-1 in unpolluted environment. Values for other allergens are scarce in literature; only one quantitative assessment exists for allergen groups Phl p 1, 2 and 4. The extractible lipid fraction of PPPGs is estimated between 1.7-2.2% of the total PG mass. The main chemical families of lipids reported in PPPGs are: alkanes, alkenes, alcohols, saturated and unsaturated fatty acids, di- and tri-hydroxylated fatty acids, aldehydes and sterols. Several lipid compounds with potential adjuvant effects on allergy have been specifically quantified in PPPGs: E2-like prostaglandin (PGE2), B4-like leukotriene (LTB4), unsaturated fatty acids (linoleic and linolenic acids and their hydroxylated derivatives), adenosine, vitamins and phenolic compounds. Some other biochemical characteristics such as NAD(P)H oxidase, protease activity and pollen microbiome were described in the literature. The bioaccessibility in physiological conditions has not been described for most biochemicals transported by allergenic PPPGs. There is also a considerable lack of knowledge about the potential health effects of pollen constituents other than allergens. The variability of pollen composition remains also largely unknown despite its importance for plant reproduction and allergy in an environment characterized by chemical pollution, climate change and loss of biodiversity.

Japanese cedar and cypress pollinosis updated: New allergens, cross-reactivity, and treatment.

Pollen from many tree species in the Cupressaceae family is a well-known cause of seasonal allergic diseases worldwide. Japanese cedar pollinosis and Japanese cypress pollinosis, which are caused by pollen from Japanese cedar (Cryptomeria japonica) and Japanese cypress (Chamaecyparis obtusa), respectively, are the most prevalent seasonal allergic diseases in Japan. Recently, the novel major Japanese cypress allergen Cha o 3 and the homologous Japanese cedar allergen Cry j cellulase were identified, and it was shown, for the first time, that cellulase in plants is allergenic. Although the allergenic components of pollen from both species exhibit high amino acid sequence identity, their pollinosis responded differently to allergen-specific immunotherapy (ASIT) using a standardized extract of Japanese cedar pollen. Pharmacotherapy and ASIT for Japanese cedar and cypress pollinosis have advanced considerably in recent years. In particular, Japanese cedar ASIT has entered a new phase, primarily in response to the generation of updated efficacy data and the development of new formulations. In this review, we focus on both Japanese cypress and cedar pollinosis, and discuss the latest findings, newly identified causative allergens, and new treatments. To manage pollinosis symptoms during spring effectively, ASIT for both Japanese cedar and Japanese cypress pollen is considered necessary.

Relationship between Phenolic Compounds, Antioxidant Properties, and the Allergenic Protein Mal d 1 in Different Selenium-Biofortified Apple Cultivars (Malus domestica).

Notable parts of the population in Europe suffer from allergies towards apples. To address this health problem, the analysis of the interactions of relevant allergens with other substances such as phenolic compounds is of particular importance. The aim of this study was to evaluate the correlations between the total phenolic content (TPC), polyphenol oxidase (PPO) activity, antioxidant activity (AOA), and the phenolic compound profile and the content of the allergenic protein Mal d 1 in six apple cultivars. It was found that the PPO activity and the content of individual phenolic compounds had an influence on the Mal d 1 content. With regard to the important constituents, flavan-3-ols and phenolic acids, it was found that apples with a higher content of chlorogenic acid and a low content of procyanidin trimers and/or epicatechin had a lower allergenic potential. This is probably based on the reaction of phenolic compounds (when oxidized by the endogenous PPO) with proteins, thus being able to change the conformation of the (allergenic) proteins, which further corresponds to a loss of antibody recognition. When apples were additionally biofortified with selenium, the composition of the apples, with regard to TPC, phenolic profile, AOA, and PPO, was significantly affected. Consequently, this innovative agronomic practice seems to be promising for reducing the allergenic potential of apples.

Fluorescent Immunolocalization of Arabinogalactan Proteins and Pectins in the Cell Wall of Plant Tissues.

Plant development involves constant adjustments of the cell wall composition and structure in response to both internal and external stimuli. Cell walls are composed of cellulose and non-cellulosic polysaccharides together with proteins, phenolic compounds and water. 90% of the cell wall is composed of polysaccharides (e.g., pectins) and arabinogalactan proteins (AGPs). The fluorescent immunolocalization of specific glycan epitopes in plant histological sections remains a key tool to uncover remodeling of wall polysaccharide networks, structure and components. Here, we report an optimized fluorescent immunolocalization procedure to detect glycan epitopes from AGPs and pectins in plant tissues. Paraformaldehyde/glutaraldehyde fixation was used along with LR-White embedding of the plant samples, allowing for a better preservation of the tissue structure and composition. Thin sections of the embedded samples obtained with an ultra-microtome were used for immunolocalization with specific antibodies. This technique offers great resolution, high specificity, and the chance to detect multiple glycan epitopes in the same sample. This technique allows subcellular localization of glycans and detects their level of accumulation in the cell wall. It also permits the determination of spatio-temporal patterns of AGP and pectin distribution during developmental processes. The use of this tool may ultimately guide research directions and link glycans to specific functions in plants. Furthermore, the information obtained can complement biochemical and gene expression studies.

Crystal structure of timothy grass allergen Phl p 12.0101 reveals an unusual profilin dimer.

Timothy grass pollen is a source of potent allergens. Among them, Phl p 1 and Phl p 5 are thought to be the most important, as a majority of timothy grass-allergic individuals have IgE antibodies directed against these two allergens. The profilin from timothy grass (Phl p 12) has been registered as a minor allergen, with up to 35% of individuals in populations of grass pollen allergic patients showing IgE binding to Phl p 12. Profilins are primarily minor allergens and are known for a high likelihood of co-sensitization as well as cross-reactivity situations caused by their sequence and structure similarity. The crystal structure of Phl p 12.0101 was determined and it revealed that this allergen may form an unusual dimer not previously observed among any profilins. For example, the Phl p 12 dimer has a completely different geometry and interface when compared with the latex profilin (Hev b 8) dimer that has its crystal structure determined. The structure of Phl p 12.0101 is described in the context of allergenic sensitization and allergy diagnostics. Moreover, the structure of the Phl p 12.0101 dimer is discussed, taking into account the production of recombinant allergens and their storage.

Identification of peroxidase-1 and beta-glucosidase as cross-reactive wheat allergens in grass pollen-related wheat allergy.

BACKGROUND: Some patients with wheat-dependent exercise-induced anaphylaxis (WDEIA) or wheat allergy showed negative ω-5 gliadin-specific IgE test and high level of grass pollen-specific IgE. It was presumed that these patients developed allergic reaction upon cross-reaction of their IgE antibodies raised against grass pollen allergens to wheat allergens. This study aimed to clarify clinical characteristics and wheat allergens of this phenotype of WDEIA/wheat allergy, which were tentatively diagnosed as grass pollen-related wheat allergy (GPWA). METHODS: A total of six patients with GPWA were enrolled, and controls were 17 patients with grass pollen allergy but no episode of wheat allergy, and 29 patients with other wheat allergies: 18 with conventional WDEIA and 11 with hydrolyzed wheat protein allergy. Sensitization to wheat proteins was determined by basophil activation test (BAT). IgE-binding proteins in wheat flour were identified by immunoblotting followed by mass spectrometry. Wheat allergen-specific IgE tests were established by CAP-FEIA system. RESULTS: All the six patients with GPWA were sensitized to water-soluble wheat proteins in BAT and IgE-immunoblotting, and peroxidase-1 (35 kDa) and beta-glucosidase (60 kDa) were identified as specific IgE-binding wheat proteins. The binding of patient IgE to these proteins was inhibited by pre-incubation of patient sera with grass pollen. The peroxidase-1- and beta-glucosidase-specific IgE tests identified three and four of six patients with GPWA, respectively, but only two of 29 controls, indicating high specificity of these tests. CONCLUSIONS: Peroxidase-1 and beta-glucosidase are specific wheat allergens for GPWA among grass pollen allergy and other types of wheat-induced food allergies.

The Role of Lipid Transfer Proteins as Food and Pollen Allergens Outside the Mediterranean Area.

PURPOSE OF REVIEW: To provide an overview of the prevalence and clinical manifestation of non-specific lipid transfer proteins (LTP)-mediated allergies outside the Mediterranean area and to address potential reasons for the different geographical significance of LTP-driven allergies. RECENT FINDINGS: LTPs are major allergens in the Mediterranean area, which frequently can elicit severe reactions. Pru p 3 the LTP from peach is reported as genuine allergen and is considered a prototypic marker for LTP-mediated allergies. However, both food and pollen LTP allergies exist outside the Mediterranean area, but with lower clinical significance, different immunogenicity, and less clarified role. Evidence has been reported that in areas with high exposure to pollen, in particular to mugwort, pollen-derived LTPs can act as a primary sensitizer to trigger secondary food allergies. Co-sensitization to unrelated allergens might be causative for less severe reactions in response to LTPs. However, the reason for the geographical different sensitization patterns to LTPs remains unclear.

Cloning, expression and immunological characterisation of Coc n 1, the first major allergen from Coconut pollen.

Coconut pollen has been documented to be a major contributor to the aeroallergen load in India, causing respiratory allergy in a large cohort of susceptible individuals. Here, we report the identification of the first major allergen from Coconut pollen, Coc n 1. The full-length sequence of the allergen was determined from previously identified peptides and overexpressed in E. coli. Recombinant Coc n 1 folded into a trimer and was found to possess allergenicity equivalent to its natural counterpart. Proteolytic processing of Coc n 1 led to the formation of an immunodominant ∼20 kDa C-terminal subunit and the site of cleavage was determined by amino acid microsequencing. Five linear IgE binding epitopes were predicted and mapped on the homology modelled structure of Coc n 1. Amongst three immunodominant epitopes, two were present towards the C-terminal end. Coc n 1 was found to belong to the highly diverse cupin superfamily and mimics its structure with known 7S globulin or vicilin allergens but lacks sequence similarity. Using sequence similarity networks, Coc n 1 clustered as a separate group containing unannotated cupin domain proteins and did not include known vicilin allergens except Gly m Bd 28 kDa, a Soybean major allergen. 7S globulins are major storage proteins and food allergens, but presence of such protein in pollen grains is reported for the first time. Further study on Coc n 1 may provide insights into its function in pollen grains and also in the development of immunotherapy to Coconut pollen allergy.

The Impact of Phosphorus on Plant Immunity.

Phosphorus (P) is the second most essential macronutrient in terms of limiting plant growth. The genes involved in P acquisition, transport, storage, utilization and respective regulation have been extensively studied. In addition, significant attention has been given to the crosstalk between P and other environmental stresses. In this review, we summarize recent discoveries pertaining to the emerging function of P in plant immunity. The roles of external soil P availability, internal cellular P in plants, P starvation signaling machinery and phosphate transporters in biotic interactions are discussed. We also highlight the impact of several phytohormones on the signaling convergence between cellular P and immune responses. This information may serve as a foundation for dissecting the molecular interaction between nutrient responses and plant immunity.

Allergome-wide peptide microarrays enable epitope deconvolution in allergen-specific immunotherapy.

BACKGROUND: The interaction of allergens and allergen-specific IgE initiates the allergic cascade after crosslinking of receptors on effector cells. Antibodies of other isotypes may modulate such a reaction. Receptor crosslinking requires binding of antibodies to multiple epitopes on the allergen. Limited information is available on the complexity of the epitope structure of most allergens. OBJECTIVES: We sought to allow description of the complexity of IgE, IgG4, and IgG epitope recognition at a global, allergome-wide level during allergen-specific immunotherapy (AIT). METHODS: We generated an allergome-wide microarray comprising 731 allergens in the form of more than 172,000 overlapping 16-mer peptides. Allergen recognition by IgE, IgG4, and IgG was examined in serum samples collected from subjects undergoing AIT against pollen allergy. RESULTS: Extensive induction of linear peptide-specific Phl p 1- and Bet v 1-specific humoral immunity was demonstrated in subjects undergoing a 3-year-long AIT against grass and birch pollen allergy, respectively. Epitope profiles differed between subjects but were largely established already after 1 year of AIT, suggesting that dominant allergen-specific antibody clones remained as important contributors to humoral immunity following their initial establishment during the early phase of AIT. Complex, subject-specific patterns of allergen isoform and group cross-reactivities in the repertoires were observed, patterns that may indicate different levels of protection against different allergen sources. CONCLUSIONS: The study highlights the complexity and subject-specific nature of allergen epitopes recognized following AIT. We envisage that epitope deconvolution will be an important aspect of future efforts to describe and analyze the outcomes of AIT in a personalized manner.

Evaluation of two commercial peach extracts for skin prick testing in the diagnosis of hypersensitivity to lipid transfer protein. A multicenter study.

Summary: The clinical usefulness of two commercial peach extracts for SPT (by Lofarma SpA and ALK-Abellò, respectively) was compared in a multicenter study carried out in Italy. Peach allergic patients were tested with the two extracts in parallel and underwent the detection of IgE specific for all three peach allergens currently available (Pru p1, Pru p3, and Pru p4, respectively). The two extracts were almost identical in terms of sensitivity and specificity, being able to detect virtually all patients sensitized to stable peach allergens (lipid transfer protein (LTP) and, presumably, peamaclein) but scoring negative in patients exclusively sensitive to labile allergens (either PR-10 and/or profilin). Thus, the two extracts represent an excellent tool to carry out a preliminary component-resolved diagnosis of peach allergy at the first patient visit.

Major latex protein-like encoding genes contribute to Rhizoctonia solani defense responses in sugar beet.

Sugar beets are attacked by several pathogens that cause root damages. Rhizoctonia (Greek for "root killer") is one of them. Rhizoctonia root rot has become an increasing problem for sugar beet production and to decrease yield losses agronomical measures are adopted. Here, two partially resistant and two susceptible sugar beet genotypes were used for transcriptome analysis to discover new defense genes to this fungal disease, information to be implemented in molecular resistance breeding. Among 217 transcripts with increased expression at 2 days post-infection (dpi), three resistance-like genes were found. These genes were not significantly elevated at 5 dpi, a time point when increased expression of three Bet v I/Major latex protein (MLP) homologous genes BvMLP1, BvMLP2 and BvML3 was observed in the partially resistant genotypes. Quantitative RT-PCR analysis on diseased sugar beet seedlings validated the activity of BvMLP1 and BvMLP3 observed in the transcriptome during challenge by R. solani. The three BvMLP genes were cloned and overexpressed in Arabidopsis thaliana to further dissect their individual contribution. Transgenic plants were also compared to T-DNA mutants of orthologous MLP genes. Plants overexpressing BvMLP1 and BvMLP3 showed significantly less infection whereas additive effects were seen on Atmlp1/Atmlp3 double mutants. The data suggest that BvMLP1 and BvMLP3 may contribute to the reduction of the Rhizoctonia root rot disease in sugar beet. Impact on the defense reaction from other differential expressed genes observed in the study is discussed.