Ragweed Allergen Research Articles

Fungi-sensitized individuals have unique profiles where Alt a 1 dominates promoting response to grass, ragweed and cat allergens.

The aim of our work was to determine comprehensively the sensitization profile of patients hypersensitive to fungal allergenic components in the Ukrainian population, identifying features of their co-sensitization to allergens of other groups and establishing potential relationships between causative allergens and their ability to provoke this hypersensitivity. A set of programs was developed using Python and R programming languages, implementing the K-means++ clustering method. Bayesian networks were constructed based on the created clusters, allowing for the assessment of the probabilistic interplay of allergen molecules in the sensitization process of patients. It was found that patients sensitive to fungi are polysensitized, with 84.77% of them having unique allergological profiles, comprising from 2 to several dozen allergens from different groups. The immune response to Alt a 1 may act as the primary trigger for sensitization to other allergens and may contribute to a high probability of developing sensitivity to grasses (primarily to Phl p 2), ragweed extract, and the Amb a 1 pectate lyase, as well as to pectate lyase Cry j 1 and cat allergen Fel d 1. Individuals polysensitized to molecular components of fungi were often sensitive to such cross-reactive molecules as lipocalins Fel d 4 and Can f 6, as well. Sensitivity to Ambrosia extract which dominated in the development of sensitization to ragweed pollen indicating the importance of different allergenic components of this plant's pollen. This hypothesis, along with the assumption that Phl p 2 may be the main trigger for sensitivity to grasses in patients with Alternaria allergy, requires further clinical investigation.

Non-Specific Lipid Transfer Protein Amb a 6 Is a Source-Specific Important Allergenic Molecule in Ragweed Pollen.

Pollen from common ragweed is an important allergen source worldwide and especially in western and southern Romania. More than 100 million patients suffer from symptoms of respiratory allergy (e.g., rhinitis, asthma) to ragweed pollen. Among the eleven characterized allergens, Amb a 6 is a non-specific lipid transfer protein (nsLTP). nsLTPs are structurally stable proteins in pollen and food from different unrelated plants capable of inducing severe reactions. The goal of this study was to produce Amb a 6 as a recombinant and structurally folded protein (rAmb a 6) and to characterize its physicochemical and immunological features. rAmb a 6 was expressed in Spodoptera frugiperda Sf9 cells as a secreted protein and characterized by mass spectrometry and circular dichroism (CD) spectroscopy regarding molecular mass and fold, respectively. The IgE-binding frequency towards the purified protein was evaluated using sera from 150 clinically well-characterized ragweed-allergic patients. The allergenic activities of rAmb a 6 and the nsLTP from the weed Parietaria judaica (Par j 2) were evaluated in basophil activation assays. rAmb a 6-specific IgE reactivity was associated with clinical features. Pure rAmb a 6 was obtained by insect cell expression. Its deduced molecular weight corresponded to that determined by mass spectrometry (i.e., 10,963 Da). rAmb a 6 formed oligomers as determined by SDS-PAGE under non-reducing conditions. According to multiple sequence comparisons, Amb a 6 was a distinct nsLTP with less than 40% sequence identity to currently known plant nsLTP allergens, except for nsLTP from Helianthus (i.e., 52%). rAmb a 6 is an important ragweed allergen recognized by 30% of ragweed pollen allergic patients. For certain patients, rAmb a 6-specific IgE levels were higher than those specific for the major ragweed allergen Amb a 1 and analysis also showed a higher allergenic activity in the basophil activation test. rAmb a 6-positive patients suffered mainly from respiratory symptoms. The assumption that Amb a 6 is a source-specific ragweed allergen is supported by the finding that none of the patients showing rAmb a 6-induced basophil activation reacted with Par j 2 and only one rAmb a 6-sensitized patient had a history of plant food allergy. Immunization of rabbits with rAmb a 6 induced IgG antibodies which strongly inhibited IgE binding to rAmb a 6. Our results demonstrate that Amb a 6 is an important source-specific ragweed pollen allergen that should be considered for diagnosis and allergen-specific immunotherapy of ragweed pollen allergy.

Heterogenous Induction of Blocking Antibodies against Ragweed Allergen Molecules by Allergen Extract-Based Immunotherapy Vaccines.

Currently, allergen-specific immunotherapy (AIT) for ragweed allergy is still based on natural allergen extracts. This study aimed to analyse the ability of four commercially available AIT vaccines (CLUSTOID, TYRO-SIT, POLLINEX Quattro Plus and Diater Depot) regarding their ability to induce IgG antibodies against ragweed pollen allergens in rabbits. Accordingly, the IgG reactivity of AIT-induced rabbit sera was tested for ten different ragweed pollen allergens (Amb a 1, 3, 4, 5, 6, 8, 9, 10, 11 and 12) by an ELISA. Furthermore, the ability of rabbit AIT-specific sera to block allergic patients' IgE binding to relevant ragweed allergens (Amb a 1, 4, 6, 8 and 11) and to inhibit allergen-induced basophil activation was evaluated by an IgE inhibition ELISA and a mediator release assay. Only two AIT vaccines (Diater Depot > CLUSTOID) induced relevant IgG antibody levels to the major ragweed allergen Amb a 1. The IgG responses induced by the AIT vaccines against the other ragweed allergens were low and highly heterogeneous. Interestingly, the kinetics of IgG responses were different among the AIT vaccines and even within one AIT vaccine (Diater Depot) for Amb a 1 (long-lasting) versus Amb a 8 and Amb a 11 (short-lived). This could be due to variations in allergen contents, the immunogenicity of the allergens, and different immunization protocols. The IgE inhibition experiments showed that rabbit AIT-specific sera containing high allergen-specific IgG levels were able to inhibit patients' IgE binding and prevent the mediator release with Diater Depot. The high levels of allergen-specific IgG levels were associated with their ability to prevent the recognition of allergens by patients' IgE and allergen-induced basophil activation, indicating that the measurement of allergen-induced IgG could be a useful surrogate marker for the immunological efficacy of vaccines. Accordingly, the results of our study may be helpful for the selection of personalized AIT vaccination strategies for ragweed-allergic patients.

Insect Cell-Expressed Major Ragweed Allergen Amb a 1.01 Exhibits Similar Allergenic Properties to Its Natural Counterpart from Common Ragweed Pollen.

Common ragweed pollen allergy has become a health burden worldwide. One of the major allergens in ragweed allergy is Amb a 1, which is responsible for over 90% of the IgE response in ragweed-allergic patients. The major allergen isoform Amb a 1.01 is the most allergenic isoform in ragweed pollen. So far, no recombinant Amb a 1.01 with similar allergenic properties to its natural counterpart (nAmb a 1.01) has been produced. Hence, this study aimed to produce a recombinant Amb a 1.01 with similar properties to the natural isoform for improved ragweed allergy management. Amb a 1.01 was expressed in insect cells using a codon-optimized DNA construct with a removable N-terminal His-Tag (rAmb a 1.01). The recombinant protein was purified by affinity chromatography and physicochemically characterized. The rAmb a 1.01 was compared to nAmb a 1.01 in terms of the IgE binding (enzyme-linked immunosorbent assay (ELISA), immunoblot) and allergenic activity (mediator release assay) in well-characterized ragweed-allergic patients. The rAmb a 1.01 exhibited similar IgE reactivity to nAmb a 1.01 in different IgE-binding assays (i.e., IgE immunoblot, ELISA, quantitative ImmunoCAP inhibition measurements). Furthermore, the rAmb a 1.01 showed comparable dose-dependent allergenic activity to nAmb a 1.01 regarding basophil activation. Overall, the results showed the successful expression of an rAmb a 1.01 with comparable characteristics to the corresponding natural isoform. Our findings provide the basis for an improvement in ragweed allergy research, diagnosis, and immunotherapy.

Behavior of Autumn Airborne Ragweed Pollen and Its Size-Segregated Allergens (Amb a 1): A study in Urban Saitama, Japan

The prevalence of ragweed (Ambrosia artemisiifolia) pollinosis has been increasing worldwide. This study focused on the behavior of autumn airborne pollen and the major ragweed allergen -Amb a 1 particle in urban Saitama, Japan, in 2016. Burkard sampler results showed that the airborne ragweed pollen scattering season was from September 1st to October 9th. Over 83% of sampling events had pollen counts of over 13 grains/m3, indicating the high potential health risks of ragweed pollen in the atmosphere. The results of a surface plasmon resonance immunoassay (SPR) indicated that the average Amb a 1 count was about 16.5 pg /pollen. The airborne Amb a 1 concentration was up to 4.7 ng/m3, of which about 45% was accumulated in ultrafine particles, such as particulate matter with a diameter ≤1.1 µm (PM1.1). Although ragweed pollen was hardly observed during the October 14th–17th sampling campaign, the concentration of ambient Amb a 1 particles in PM1.1 was also determined to be 4.59 ng/m3, which could be explained by the longer scattering of fine particles in the atmosphere. Pearson correlation coefficient analysis results showed that temperature (daily, r = 0.41; event, r = 0.87) could affect the behavior of the airborne pollen counts, and ambient water-soluble ions (such as Ca2+ and NO3−) could affect Amb a 1 in PM1.1. Additionally, air mass trajectories and wind rose results indicated that air masses with long-range transportation could also influence the temporary behavior of Amb a 1 and pollen counts via the wind. Mugwort and Humulus japonicus pollens were also observed to extend pollen scattering periods. Airborne pollen and allergenic particles could be considered air pollutants, as they pose health risks and are susceptible to environmental influences.

Determination of ragweed allergen Amb a 1 distribution in aerosols using ELISA and immunogold scanning electron microscopy.

Ragweed as an invasive species in Europe has become more important for allergy sufferers in the last decade. Because pollen fractions can be found in the respirable fraction of aerosols, they can generate severe disease progressions. To obtain information about the concentration and distribution of 1 of the main ragweed allergens Ambrosia artemisiifolia 1 in the air of Vienna, PM10 and PM2.5 fine dust filters were analyzed. Standard fine dust filters used for air quality monitoring were analyzed via ELISA and immunogold scanning electron microscopy. Via ELISA it was possible to show that already at pollen season start in August a recognizably high A artemisiifolia 1 concentration can be found. In addition, the allergen concentration in the air stays comparatively high after the peak season has ended even when the pollen concentration drops to a moderate level. The immunogold electron microscopy investigation directly applied on filters shows that the allergen can be found on organic as well as on mixtures of organic and inorganic particles. Afirst semistatistical analysis of the labeled particle sizes indicates that a large number of the allergen carriers can be found within the smallest particle size range. Nevertheless, further investigations are needed to obtain enough particle counts for a significant statistical analysis. It was possible to show that reliable results can be obtained from ELISA and immunogold scanning electron microscopy directly applied on filters that are used in air qualitymonitoring sites. By adaptation of the used protocols, itshould be possible to obtain respective information aboutfurther allergens.

Suppression spontaneous and induced expression of CD203c basophils after 
 subcutaneous immunotherapy with ragweed and mugwort allergens

BACKGROUND: The basophil activation test (BAT) reflects the presence of IgE on the cell surface and can be a tool for monitoring the effectiveness of allergen-specific immunotherapy (ASIT). It is likely that spontaneous and allergen-induced BAT levels may change after effective subcutaneous immunotherapy (SCIT) and this is of clinical importance.
 AIMS: determination of basophil activation markers in patients sensitized to weed pollen during SCIT.
 MATERIALS AND METHODS: A real clinical study (2 years, 60 patients) with allergic rhinitis who underwent SCIT with weed allergens for 2 seasons. The proportion of basophils expressing CD203c spontaneously and after exposure to a trigger allergen was determined. BAT was performed before treatment, after the first and second courses of pre-season SCIT (0, 3, 15 months of treatment). The effectiveness of SCIT was assessed by the dynamics of the symptom scale (Rhinoconjunctivitis total symptom score) and the drug assessment (Daily medical score).
 RESULTS: SCIT with water-salt extracts was effective: median RTSS in patients after two courses decreased from 13.0 [10.0; 15.0] to 4.5 [3.00; 7.25] points (p 0.001), DMS score from 2.0 [2.0; 2.0] vs 1.0 [1.0; 2.0] points (p0.001).
 Spontaneous BAT remained unchanged after the first course of ASIT. Mean BAT values ​​before therapy and after the first course of SCIT did not differ (8.15 [6.10; 11.9] vs 9.0 [6.30; 12.0] %). After the second course of treatment, spontaneous BAT decreased (8.15 [6.10; 11.9] vs 6.20 [4.27; 9.00] %, 2 = 15.5, p0.001).
 Allergen-induced BAT decreased significantly from the first year of treatment. The overall change in the total group for BAT with ragweed after the first course of SCIT was 11.25 (CI 95 %: 7.10; 15.10) %, p 0.001. The overall difference in the induced ВAT test at the end of therapy reached 8.63 (CI 95: 5.30; 12.05) %, p 0.001. After the first course of therapy with mugwort allergens, the indices of induced BAT decreased by 12.0 (CI 95 %: 6.95; 17.90) %, p 0.001; after the second course, the difference from the initial values ​​was 9.67 (CI 95%: -6.85; 15.90) %, p0.001.
 CONCLUSIONS: SCIT with ragweed and mugwort allergens was effective. The change in spontaneous and induced BAT is associated with a positive effect of ASIT.

Complex IgE sensitization patterns in ragweed allergic patients: Implications for diagnosis and specific immunotherapy.

BackgroundRagweed (Ambrosia artemisiifolia) is one of the most important allergen sources, worldwide, causing severe respiratory allergic reactions in late summer and fall, in sensitized patients. Amb a 1 has been considered as the most important allergen in ragweed but 12 ragweed pollen allergens are known. The aim of our study was to investigate IgE reactivity profiles of ragweed allergic patients and to associate them with clinical symptoms.MethodsIgE sensitization profiles from clinically well‐characterized ragweed allergic patients (n = 150) were analyzed using immunoblotted ragweed pollen extract. Immunoblot inhibition experiments were performed with two Amb a 1 isoforms and CCD markers and basophil activation experiments were performed with IgE serum before and after depletion of Amb a 1‐specific IgE.ResultsBy IgE‐immunoblotting 19 different IgE reactivity patterns with and without Amb a 1‐sensitization were found. The majority of patients (>95%) suffered from rhino‐conjunctivitis, around 60% reported asthma‐like symptoms and about 25% had skin reactions. Patients with complex IgE sensitization profiles tended to have more clinical symptoms. Serum with and without Amb a 1‐specific IgE induced basophil activation.ConclusionsRagweed pollen allergic patients exhibit complex IgE reactivity profiles to ragweed allergens including Amb a 1 isoforms and cross‐reactive carbohydrates indicating the importance of Amb a 1 isoforms and additional allergens for diagnosis and allergen‐specific immunotherapy of ragweed allergy.

Evaluation of the effectiveness of subcutaneous pollen weed polen allergens

Background. In European countries, the prevalence of pollen sensitization reaches 30–40%. In Russia pollen allergy ranges from 12,7 to 38%. In the southern regions of Russia, the main one is ragweed and weed pollen. The efficacy of the subcutaneous medicinal ragweed allergen has not previously been evaluated using modern symptom scales.
 Methods. Open-label, single-center, placebo-uncontrolled study of 60 patients aged 16–55 years with allergic rhinitis in combination with bronchial asthma and without it. A 2-preseason course of AIT with a subcutaneous allergen of Ambrosia was used for patients with monosensitization and an allergen Ambrosia + Artemisia (for oligosensitized patients) and Ambrosia + a mix of pollen of causal allergen for polysensitized patients. Treatment started 4 months before and stopped 2–4 weeks before the beginning of pollination. The RTSS (the rhinoconjunctivitis total symptom score) scale was used to assess the symptoms, and the DMS (Daily medical score) was used to estimate of medical treatment.
 Results. After 1-st season of treatment, there was a rapid decrease in the RTSS to 5,0 [4,0; 9,0] compared to the initial values of 13 [10, 15] points. By the second year, the decline in RTSS amounted to 4,5 [3,0; 4,5] points (p <0,001), the total effect size by second course was 0,96 (strong effect of the intervention).
 DMS (before treatment) was 2,0 [2,0; 2,0] points, after the first and second courses — 1,0 [1,0; 2,0] points (p=0,002), with a moderate effect size of 0,67 after the first year; by the second year, the effect size is 0,75 versus baseline (p <0,01).
 Conclusion. Monotherapy with a subcutaneous ragweed allergen or its combination with other allergens show high effectiveness, starting from the first course of therapy. In the second year, the improvement continues, but the dynamics decreases. The study demonstrated a strong size of the effect of medical intervention, which cannot be explained by the placebo effect.

Ragweed sublingual immunotherapy (SLIT) tablets in allergic rhinoconjunctivitis: a systematic review and meta-analysis.

Ragweed allergen causes Allergic rhinoconjunctivitis and sublingual immunotherapy is one of the treatment modalities to desensitize allergic individuals. This systematic review assesses the effectiveness and safety of sublingual immunotherapy for allergic rhinoconjunctivitis caused due to Ragweed. The databases search was done through December 2020. English-language randomized controlled trials were included if they compared sublingual immunotherapy with placebo, pharmacotherapy, or other sublingual immunotherapy regimens, and reported clinical outcomes. The strength of the evidence for each comparison and outcome was graded based on the risk of bias, consistency, magnitude of effect, and the directness of the evidence. The searches performed according to the protocol identified 134 abstracts of which 67 were duplicates. A total of 37 full papers were therefore reviewed of which 5 were included for the final study. Participants' ages ranged from 4 to 58years. The risk of bias was low in most studies. The review suggests that sublingual immunotherapy improves rhinoconjunctivitis symptoms, with 4 of 4 studies reporting efficacy showed improvement in the symptom score of SLIT groups compared to placebo. Local reactions were frequent, but anaphylaxis was not reported in any of the studies. Serious adverse events were very few in all the studies. The overall evidence showed the effectiveness of sublingual immunotherapy for the treatment of allergic rhinoconjunctivitis with or without asthma, but high-quality studies are still needed to answer questions regarding optimal dosing strategies.

Dectin-1 Controls TSLP-Induced Th2 Response by Regulating STAT3, STAT6, and p50-RelB Activities in Dendritic Cells.

The epithelium-associated cytokine thymic stromal lymphopoietin (TSLP) can induce OX40L and CCL17 expression by myeloid dendritic cells (mDCs), which contributes to aberrant Th2-type immune responses. Herein, we report that such TSLP-induced Th2-type immune response can be effectively controlled by Dectin-1, a C-type lectin receptor expressed by mDCs. Dectin-1 stimulation induced STAT3 activation and decreased the transcriptional activity of p50-RelB, both of which resulted in reduced OX40L expression on TSLP-activated mDCs. Dectin-1 stimulation also suppressed TSLP-induced STAT6 activation, resulting in decreased expression of the Th2 chemoattractant CCL17. We further demonstrated that Dectin-1 activation was capable of suppressing ragweed allergen (Amb a 1)-specific Th2-type T cell response in allergy patients ex vivo and house dust mite allergen (Der p 1)-specific IgE response in non-human primates in vivo. Collectively, this study provides a molecular explanation of Dectin-1-mediated suppression of Th2-type inflammatory responses and suggests Dectin-1 as a target for controlling Th2-type inflammation.

Prevalence of sensitization to specific allergens in allergic patients in Beijing, China: A 7-year retrospective study.

Knowledge of the prevalence of common sensitizing allergens may aid in overall management of allergic disease in a specified area. The aim of this study was to identify and analyse the prevalence of common inhaled and food sensitizing allergens in Beijing. This was a retrospective study, analysing demographic data and serum sIgE antibody test results from 59057 outpatients who presented to Beijing TongRen Hospital, from January 2013 to December 2019. 28879 patients (48.9%) showed positive sIgE test results; with significantly more males aged under 16 years sensitized to at least one allergen than females, and most patients (53.62%) were sensitized to multiple allergens. The first inhaled sensitizing allergens was Artemisia grass (11910 (41.24%)); and the first food allergens was crab (3547 (12.28%)). For Artemisia sensitized patients, sIgE levels were mostly at level 5. The number of patients with ragweed allergy is increasing year by year. The detection rates for sIgE to Artemisia, common ragweed, and Humulus grass allergens were significantly higher in August and September. R package ggplot2 analysis, demonstrated strong correlations between tree allergens and common ragweed and Humulus grass allergens (phi coefficients = 0.50 and 0.46, respectively; both P < 0.01). The prevalence of sensitization to different allergens in Beijing showed Artemisia grass was the most commonly inhaled sensitizing allergen, and the number of patients with ragweed grass allergy was increasing by year.

Notch 2 receptor expression and reduced cytotoxicity in MAIT cells of active pulmonary TB patients.

Knowledge of the prevalence of common sensitizing allergens may aid in overall management of allergic disease in a specified area. The aim of this study was to identify and analyse the prevalence of common inhaled and food sensitizing allergens in Beijing. This was a retrospective study, analysing demographic data and serum sIgE antibody test results from 59057 outpatients who presented to Beijing TongRen Hospital, from January 2013 to December 2019. 28879 patients (48.9%) showed positive sIgE test results; with significantly more males aged under 16 years sensitized to at least one allergen than females, and most patients (53.62%) were sensitized to multiple allergens. The first inhaled sensitizing allergens was Artemisia grass (11910 (41.24%)); and the first food allergens was crab (3547 (12.28%)). For Artemisia sensitized patients, sIgE levels were mostly at level 5. The number of patients with ragweed allergy is increasing year by year. The detection rates for sIgE to Artemisia, common ragweed, and Humulus grass allergens were significantly higher in August and September. R package ggplot2 analysis, demonstrated strong correlations between tree allergens and common ragweed and Humulus grass allergens (phi coefficients = 0.50 and 0.46, respectively; both P < 0.01). The prevalence of sensitization to different allergens in Beijing showed Artemisia grass was the most commonly inhaled sensitizing allergen, and the number of patients with ragweed grass allergy was increasing by year.

Homology modeling of Amb a 4 and Amb a 6 ragweed allergens and in silico prediction of their T-cell and B-cell epitopes for peptide-based immunotherapy

Homology modeling of Amb a 4 and Amb a 6 ragweed allergens and in silico prediction of their T-cell and B-cell epitopes for peptide-based immunotherapy

Allergen diagnosis strategy: An experimental application of different methods in Guangzhou, Southern China.

Different methods have different diagnostic value under diverse clinical conditions. This study was conducted to analyze the clinical practical value of different assays in different situations and to establish the concept of an allergen diagnosis strategy.This study used Phadia CAP, Advan Sure (AdvS), Euroline Camera (ELC), and Allergy Lateral Flow Assay (ALFA) kits to detect allergen specific immunoglobulin E (sIgE) in the sera from 30 allergic asthma patients, with Phadia CAP as the gold standard.This study found high sensitivity for Dermatophagoides pteronyssinus in AdvS (92.6%), ELC (88.9%), and ALFA (85.2%), but poor sensitivity for F. domesticus in all three assays. Furthermore, the AdvS, ELC, and ALFA assays could detect 20, 21, and 8 allergens in one panel, respectively. The combination of the results showed that 93.3% and 46.7% of patients tested positive for ≥5 and ≥10 types of allergens, respectively. All patients who were tested positive for the Marine fish mixture were positive for both Shrimp and Crab. Similarly, patients who were tested positive for Chicken also tested positive for Pork. Optimal scale analysis showed a strong connection between Peanut, Soybean, Wheat, Japanese Hop, Western ragweed, and Oak allergens (Cronbach's α = 91.1%).The ELC assay was more suitable for use in pollen-rich areas, whereas the ALFA assay was easy to establish in areas with poor medical conditions. An allergen diagnosis strategy needs to be considered for use in clinical practice.

Examining sIgE-profile in patients with ambrosia allergy in Samara

It is considered that Ambrosia trifida is not widespread in Russia, thereby avoiding its threatening effects to populational health. However, there are some areas in Russia where more than half of land area is covered by Ambrosia trifida. Many such foci are found in the Samara, Orenburg regions, Tatarstan and Bashkiria. Ambrosia trifida is distributed in Central Russia, the North Caucasus region and Siberia. The objective of the study was to examine specific IgE-profile in ragweed sensitized patients in the Samara region. A clinical and allergy examination of 969 patients with allergic rhinitis was performed. Allergic diagnostics was performed by using skin testing (prick-test) with standard set of pollen, household, and epidermal allergens. The data of skin tests in patients co-sensitized with ragweed and Artemisia allowed to analyze IgE-antibodies specific to the major Ambrosia (Amb a 1, Аmb. trifida) and Artemisia (Art v 1) allergens. Patients with negative skin test for the Amdrosia artemisiifolia and clinical manifestations of seasonal allergy were assessed for level of sIgE-antibodies nAmb a 1 and Amb trif. Skin test data demonstrated that seasonal and combined forms of allergic rhinitis dominated in patients examined. In particular, positive skin test data for pollen allergens dominated (50.6%) among all allergen groups so that more than half of them belonged to weed pollen allergens. Patients challenged with skin tests for Artemisia and ragweed allergens more likely displayed positive reaction to the Artemisia allergen (71.27%) and Ambrosia artemisiifolia (50.88%). Co-sensitization (to Artemisia and ragweed) prevailed in this geographic region featured with equal rate of simultaneously detected sIgE specific to the two types of ragweed and artemisia (nAmb a 1 + nArt v 1 + Amb trif) as well as Ambrosia trifida and Artemisia (Amb trif + nArt v 1). Almost half of the patients with negative Ambrosia art. prick-test were found to bear sIgE specific to Amb trif. Further investigation will help to better understand this phenomenon and take a fresh look at diagnostics and treatment of ragweed allergy in relevant geographic area.

Stability of extracts from pollens of allergenic importance in Korea.

Background/AimsAccurate diagnosis and the effects of allergen-specific immunotherapy for pollinosis are greatly dependent on the potency and stability of the extract. This study aimed to examine factors, such as temperature and storage buffer composition, that affect the stability of allergen extracts from pollens of allergenic importance in Korea.MethodsWe prepared four pollen allergen extracts from ragweed, mugwort, Japanese hop, and sawtooth oak, which are the most important causes of seasonal rhinitis in Korea. Changes of protein and major allergen concentration were measured over 1 year by Bradford assay, two-site enzyme-linked immunosorbent assay, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis after reconstitution of the lyophilized allergen extract in various buffers and stored at room temperature (RT, 18°C to 26°C) or refrigerated (4°C).ResultsMore than 90% of the original protein concentration in all four extracts examined was detected over 1 year when 50% glycerol was added and refrigerated, whereas 57.9% to 94.5% remained in the extracts at RT. The addition of 50% glycerol to the storage buffer was found to prevent protein degradation at RT. Amb a 1, a major allergen of ragweed, was almost completely degraded in 9 weeks at RT when reconstituted in a buffer without 50% glycerol. However, 55.6% to 92.8% of Amb a 1 content was detected after 1 year of incubation at 4°C in all buffer conditions except 0.3% phenol.ConclusionsAddition of 50% glycerol as well as refrigeration was found to be important in increasing the shelf-life of allergen extracts from pollens of allergenic importance.

Detection of reactive allergens in long-distance transported pollen grains: Evidence from Ambrosia

The pollen of Ambrosia artemisiifolia (ragweed) is an important aeroallergen. The plant originated from North America and, has spread worldwide in recent decades. Air masses can transport pollen grains far from their source region and episodes of long distance transported (LDT) of ragweed pollen have been reported. The allergenic properties of LDT pollen grains can be altered by humidity, temperature and UV radiation. The aims of the study are to detect the major allergen of ragweed (Amb a 1) in the atmosphere of uninfested areas and to compare daily Ambrosia sp. pollen grains with Amb a 1 amount per m3 of air. Samples for allergen detection were collected by ChemVol® sampler. ChemVol® collects particles at 800 l/min and contains 2 impaction stages (PM > 10 μm and 10 > PM > 2.5 μm). Samples collected on to filters were obtained during a 63 day period in the year 2014. Amb a 1 was recorded on 54 days and pollen grains of ragweed on 43 days. Diurnal average Amb a 1 concentrations ranged between 0.29 and 263.3 pg per cubic meter of air and correlated positively with daily average ragweed pollen concentrations. The average seasonal Ambrosia pollen allergen potency was 2.57 pg Amb a 1/pollen. The air mass movement was simulated using the HYbrid Single-particle Lagrangian Integrated Trajectory (HYSPLIT) model and back-trajectory paths were computed for four episodes. The findings suggest that ragweed pollen allergens still survive after at least 48 h during transport over long distances from allochthonous regions and are therefore a potential trigger of allergic reactions even in areas where ragweed is not widely dispersed.

Subpollen particles are rich carriers of major short ragweed allergens and NADH dehydrogenases: quantitative proteomic and allergomic study.

Short ragweed (Ambrosia artemisiifolia) allergies affect more than 36 million people annually. Ragweed pollen grains release subpollen particles (SPP) of respirable size upon hydration or a change in air electrical conditions. The aim of this study was to characterize the proteomes and allergomes of short ragweed SPP and total pollen protein extract (TOT), and compare their effects with those of standard aqueous pollen protein extract (APE) using sera from short ragweed pollen-sensitized patients. Quantitative 2D gel-based and shotgun proteomics, 1D and 2D immunoblotting, and quantitative ELISA were applied. Novel SPP extraction and preparation protocols enabled appropriate sample preparation and further downstream analysis by quantitative proteomics. The SPP fraction contained the highest proportion (94%) of the allergome, with the largest quantities of the minor Amb a 4 and major Amb a 1 allergens, and as unique, NADH dehydrogenases. APE was the richest in Amb a 6, Amb a 5 and Amb a 3, and TOT fraction was the richest in the Amb a 8 allergens (89% and 83% of allergome, respectively). Allergenic potency correlated well among the three fractions tested, with 1D immunoblots demonstrating a slight predominance of IgE reactivity to SPP compared to TOT and APE. However, the strongest IgE binding in ELISA was noted against APE. New allergenic candidates, phosphoglycerate mutase and phosphoglucomutase, were identified in all the three pollen fractions. Enolase, UTP-glucose-1-phosphate uridylyltransferase and polygalacturonase were observed in SPP and TOT fractions as novel allergens of the short ragweed pollen, as previously described. We demonstrated that the complete major (Amb a 1 and 11) and almost all minor (Amb a 3, 4, 5, 6, 8 and 9) short ragweed pollen allergen repertoire as well as NADH oxidases are present in SPP, highlighting an important role for SPP in allergic sensitization to short ragweed.

Sequence conservation predicts T cell reactivity against ragweed allergens

Ragweed is a major cause of seasonal allergy, affecting millions of people worldwide. Several allergens have been defined based on IgE reactivity, but their relative immunogenicity in terms of T cell responses has not been studied. We comprehensively characterized T cell responses from atopic, ragweed-allergic subjects to Amb a 1, Amb a 3, Amb a 4, Amb a 5, Amb a 6, Amb a 8, Amb a 9, Amb a 10, Amb a 11, and Amb p 5 and examined their correlation with serological reactivity and sequence conservation in other allergens. Peripheral blood mononuclear cells (PBMCs) from donors positive for IgE towards ragweed extracts after in vitro expansion for secretion of IL-5 (a representative Th2 cytokine) and IFN-γ (Th1) in response to a panel of overlapping peptides spanning the above-listed allergens were assessed. Three previously identified dominant T cell epitopes (Amb a 1 176-191, 200-215, and 344-359) were confirmed, and three novel dominant epitopes (Amb a 1 280-295, 304-319, and 320-335) were identified. Amb a 1, the dominant IgE allergen, was also the dominant T cell allergen, but dominance patterns for T cell and IgE responses for the other ragweed allergens did not correlate. Dominance for T cell responses correlated with conservation of ragweed epitopes with sequences of other well-known allergens. These results provide the first assessment of the hierarchy of T cell reactivity in ragweed allergens, which is distinct from that observed for IgE reactivity and influenced by T cell epitope sequence conservation. The results suggest that ragweed allergens associated with lesser IgE reactivity and significant T cell reactivity may be targeted for T cell immunotherapy, and further support the development of immunotherapies against epitopes conserved across species to generate broad reactivity against many common allergens.