A novel grass pollen allergen mimotope identified by phage display peptide library inhibits allergen-human IgE antibody interaction

The aim of this study was to investigate the molecular basis of human IgE-allergen interaction by screening a phage-displayed peptide library with an allergen-specific human IgE-mimicking monoclonal antibody (mAb). A mAb that reacted with major grass pollen allergens was successfully identified and shown to inhibit human IgE-allergen interaction. Biopanning of a phage-displayed random peptide library with this mAb yielded a 12 amino acid long mimotope. A synthetic peptide based on this 12-mer mimotope inhibited mAb and human IgE binding to grass pollen extracts. Our results indicate that such synthetic peptide mimotopes of allergens have potential as novel therapeutic agents. © 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

Constructing a 7-day Ahead Forecast Model for Grass Pollen at North London, United Kingdom

A number of media outlets now issue medium-range (~7 day) weather forecasts on a regular basis. It is therefore logical that aerobiologists should attempt to produce medium-range forecasts for allergenic pollen that cover the same time period as the weather forecasts. The objective of this study is to construct a medium-range (< 7 day) forecast model for grass pollen at north London. The forecast models were produced using regression analysis based on grass pollen and meteorological data from 1990-1999 and tested on data from 2000 and 2002. The modelling process was improved by dividing the grass pollen season into three periods; the pre-peak, peak and post peak periods of grass pollen release. The forecast consisted of five regression models. Two simple linear regression models predicting the start and end date of the peak period, and three multiple regression models forecasting daily average grass pollen counts in the pre-peak, peak and post-peak periods. Overall the forecast models achieved 62% accuracy in 2000 and 47% in 2002, reflecting the fact that the 2002 grass pollen season was of a higher magnitude than any of the other seasons included in the analysis. This study has the potential to make a notable contribution to the field of aerobiology. Winter averages of the North Atlantic Oscillation were used to predict certain characteristics of the grass pollen season, which presents an important advance in aerobiological work. The ability to predict allergenic pollen counts for a period between five and seven days will benefit allergy sufferers. Furthermore, medium-range forecasts for allergenic pollen will be of assistance to the medical profession, including allergists planning treatment and physicians scheduling clinical trials.

Personal Exposure to Grass Pollen: Relating Inhaled Dose to Background Concentration

Background Very few studies on human exposure to allergenic pollen have been conducted using direct methods, with background concentrations measured at city center monitoring stations typically taken as a proxy for exposure despite the inhomogeneous nature of atmospheric pollen concentrations. A 2003 World Health Organization report highlighted the need for an improved understanding of the relation between monitoring station data and actual exposure. Objective To investigate the relation between grass pollen dose and background concentrations measured at a monitoring station, to assess the fidelity of monitoring station data as a qualitative proxy for dose, and to evaluate the ratio of dose rate to background concentration. Methods Grass pollen dose data were collected in Aarhus, Denmark, in an area where grass pollen sources were prevalent, using Nasal Air Samplers. Sample collection lasted for approximately 25 to 30 minutes and was performed at 2-hour intervals from noon to midevening under moderate exercise by 2 individuals. Results A median ratio of dose rate to background concentration of 0.018 was recorded, with higher ratio values frequently occurring at 12 to 2 pm, the time of day when grass species likely to be present in the area are expected to flower. From 4 to 8 pm, dose rate and background concentration data were found to be strongly and significantly correlated (rs = 0.81). Averaged dose rate and background concentration data showed opposing temporal trends. Conclusion Where local emissions are not a factor, background concentration data constitute a good quantitative proxy for inhaled dose. The present ratio of dose rate to background concentration may aid the study of dose–response relations.

Variation of the Group 5 Grass Pollen Allergen Content of Airborne Pollen in Relation to Geographical Location and Time in Season

Allergies to grass pollen are the number one cause of outdoor hay fever. The human immune system reacts with symptoms to allergens from pollen. Objective: We investigated the natural variability in release of the major group 5 allergen from grass pollen across Europe. Methods: Airborne pollen and allergens were simultaneously collected daily with a volumetric spore trap and a high-volume cascade impactor at 10 sites across Europe for 3 consecutive years. Group 5 allergen was determined with a Phl p 5 specific ELISA in two fractions of ambient air: Particulate Matter (PM) >10μm and 10μm>PM>2.5μm. Mediator release by ambient air was determined in FcεR1-humanized basophils. Origin of pollen was modeled and condensed to pollen potency maps. Results: On average grass pollen released 2.3 pg Phl p 5/pollen. Allergen release per pollen (potency) varied substantially, ranging from 0 to 9 pg Phl p 5/pollen (5 to 95% percentile). The main variation was locally day-to-day. Average potency maps across Europe varied between years. Mediator release from basophilic granulocytes correlated better with allergen/m3 (r2=0.80, p<0.001) than with pollen/m3 (r2=0.61, p<0.001). In addition, pollen released different amounts of allergen in the nonpollen bearing fraction of ambient air depending on humidity. Conclusion: Across Europe, the same amount of pollen released substantially different amounts of group 5 grass pollen allergen. This variation in allergen release is on top of variations in pollen counts. Molecular aerobiology, i.e. determining allergen in ambient air, may be a valuable addition to pollen counting.

Anti-idiotypic Fab Fragments Image a Conserved N-terminal Epitope Patch of Grass Pollen Allergen Phl p 1

BACKGROUND AND AIMS: Naturally occurring anti-idiotypic antibodies structurally mimic the original antibody epitope. Anti-idiotypes, therefore, are interesting tools for the portrayal of conformational B-cell epitopes of allergens. In this study we used this strategy particularly for major timothy grass pollen (Phleum pratense) allergen Phl p 1. METHODS AND RESULTS: We used a combinatorial phage display library constructed from the peripheral IgG repertoire of a grass pollen allergic patient which was supposed to contain anti-idiotypic Fab specificities. Using purified anti-Phl p 1 IgG for biopanning, several Fab displaying phage clones could be isolated. 100 amplified colonies were screened for their binding capacity to anti-Phl p 1-specific antibodies, finally resulting in four distinct Fab clones according to sequence analysis. Interestingly, heavy chains of all clones derived from the same germ line sequence and showed high homology in their CDRs. Projecting their sequence information on the surface of the natural allergen Phl p 1 (PDB ID: 1N10) indicated matches on the N-terminal domain of the homo-dimeric allergen, including the bridging region between the two monomers. The resulting epitope patches were formed by spatially distant sections of the primary allergen sequence. CONCLUSION: In this study we report that anti-idiotypic specificities towards anti-Phl p 1 IgG, selected from a Fab library of a grass pollen allergic patient, mimic a conformational epitope patch being distinct from a previously reported IgE epitope area.

IgE+ B cells are scarce, but allergen-specific B cells with a memory phenotype circulate in patients with allergic rhinitis

Background Despite the critical role of immunoglobulin E (IgE) in allergy, circulating IgE+ B cells are scarce. Here, we describe in patients with allergic rhinitis B cells with a memory phenotype responding to a prototypic aeroallergen. Methods Fifteen allergic rhinitis patients with grass pollen allergy and 13 control subjects were examined. Blood mononuclear cells stained with carboxyfluorescein diacetate succinimidyl ester (CFSE) were cultured with Bahia grass pollen. Proliferation and phenotype were assessed by multicolour flow cytometry. Results In blood of allergic rhinitis patients with high serum IgE to grass pollen, most IgEhi cells were CD123+ HLA-DR- basophils, with IgE for the major pollen allergen (Pas n 1). Both B and T cells from pollen-allergic donors showed higher proliferation to grass pollen than nonallergic donors (P = 0.002, and 0.010, respectively), whereas responses to vaccine antigens and mitogen did not differ between groups. Allergen-driven B cells that divided rapidly (CD19mid CD3- CFSElo) showed higher CD27 (P = 0.008) and lower CD19 (P = 0.004) and CD20 (P = 0.004) expression than B cells that were slow to respond to allergen (CD19hi CD3- CFSEmid). Moreover, rapidly dividing allergen-driven B cells (CD19mid CFSElo CD27hi) showed higher expression of the plasmablast marker CD38 compared with B cells (CD19hi CFSEmid CD27lo) that were slow to divide. Conclusion Patients with pollen allergy but not control donors have a population of circulating allergen-specific B cells with the phenotype and functional properties of adaptive memory B-cell responses. These cells could provide precursors for allergen-specific IgE production upon allergen re-exposure. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Regional and seasonal variation in airborne grass pollen levels between cities of Australia and New Zealand

Although grass pollen is widely regarded as the major outdoor aeroallergen source in Australia and New Zealand (NZ), no assemblage of airborne pollen data for the region has been previously compiled. Grass pollen count data collected at 14 urban sites in Australia and NZ over periods ranging from 1 to 17 years were acquired, assembled and compared, revealing considerable spatiotemporal variability. Although direct comparison between these data is problematic due to methodological differences between monitoring sites, the following patterns are apparent. Grass pollen seasons tended to have more than one peak from tropics to latitudes of 37°S and single peaks at sites south of this latitude. A longer grass pollen season was therefore found at sites below 37°S, driven by later seasonal end dates for grass growth and flowering. Daily pollen counts increased with latitude; subtropical regions had seasons of both high intensity and long duration. At higher latitude sites, the single springtime grass pollen peak is potentially due to a cooler growing season and a predominance of pollen from C3 grasses. The multiple peaks at lower latitude sites may be due to a warmer season and the predominance of pollen from C4 grasses. Prevalence and duration of seasonal allergies may reflect the differing pollen seasons across Australia and NZ. It must be emphasized that these findings are tentative due to limitations in the available data, reinforcing the need to implement standardized pollen-monitoring methods across Australasia. Furthermore, spatiotemporal differences in grass pollen counts indicate that local, current, standardized pollen monitoring would assist with the management of pollen allergen exposure for patients at risk of allergic rhinitis and asthma. © 2015 Springer Science+Business Media Dordrecht

Differences in grass pollen allergen exposure across Australia

Objective Allergic rhinitis and allergic asthma are important chronic diseases posing serious public health issues in Australia with associated medical, economic, and societal burdens. Pollen are significant sources of clinically relevant outdoor aeroallergens, recognised as both a major trigger for, and cause of, allergic respiratory diseases. This study aimed to provide a national, and indeed international, perspective on the state of Australian pollen data using a large representative sample. Methods Atmospheric grass pollen concentration is examined over a number of years within the period 1995 to 2013 for Brisbane, Canberra, Darwin, Hobart, Melbourne, and Sydney, including determination of the clinical' grass pollen season and grass pollen peak. Results The results of this study describe, for the first time, a striking spatial and temporal variability in grass pollen seasons in Australia, with important implications for clinicians and public health professionals, and the Australian grass pollen-allergic community. Conclusions These results demonstrate that static pollen calendars are of limited utility and in some cases misleading. This study also highlights significant deficiencies and limitations in the existing Australian pollen monitoring and data. Implications: Establishment of an Australian national pollen monitoring network would help facilitate advances in the clinical and public health management of the millions of Australians with asthma and allergic rhinitis.

A rapid and efficient two-step gel electrophoresis method for the purification of major rye grass pollen allergens

Purified proteins are mandatory for molecular, immunological and cellular studies. However, purification of proteins from complex mixtures requires specialised chromatography methods (i.e., gel filtration, ion exchange, etc.) using fast protein liquid chromatography (FPLC) or high-performance liquid chromatography (HPLC) systems. Such systems are expensive and certain proteins require two or more different steps for sufficient purity and generally result in low recovery. The aim of this study was to develop a rapid, inexpensive and efficient gel-electrophoresis-based protein purification method using basic and readily available laboratory equipment. We have used crude rye grass pollen extract to purify the major allergens Lol p 1 and Lol p 5 as the model protein candidates. Total proteins were resolved on large primary gel and Coomassie Brilliant Blue (CBB)-stained Lol p 1/5 allergens were excised and purified on a secondary "mini"-gel. Purified proteins were extracted from unstained separating gels and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses. Silver-stained SDS-PAGE gels resolved pure proteins (i.e., 875 μg of Lol p 1 recovered from a 8 mg crude starting material) while immunoblot analysis confirmed immunological reactivity of the purified proteins. Such a purification method is rapid, inexpensive, and efficient in generating proteins of sufficient purity for use in monoclonal antibody (mAb) production, protein sequencing and general molecular, immunological, and cellular studies.

A 30-Day-Ahead Forecast Model for Grass Pollen in North London, United Kingdom

A 30-day ahead forecast method has been developed for grass pollen at north London. The total period of the grass pollen season is covered by eight multiple regression models, each covering a 10-day period running consecutively from 21st May to 8th August. This means that three models were used for each 30-day forecast. The forecast models were produced using grass pollen and environmental data from 1961-1999 and tested on data from 2000 and 2002. Model accuracy was judged in two ways: the number of times the forecast model was able to successfully predict the severity (relative to the 1961-1999 dataset as a whole) of grass pollen counts in each of the eight forecast periods on a scale of one to four; and the number of times the forecast model was able to predict whether grass pollen counts were higher or lower than the mean. The models achieved 62.5% accuracy in both assessment years when predicting the relative severity of grass pollen counts on a scale of one to four, which equates to six of the eight 10-day periods being forecast correctly. The models attained 87.5% and 100% accuracy in 2000 and 2002 respectively when predicting whether grass pollen counts would be higher or lower than the mean. Attempting to predict pollen counts during distinct 10-day periods throughout the grass pollen season is a novel approach. The models also employed original methodology in the use of winter averages of the North Atlantic Oscillation to forecast 10-day means of allergenic pollen counts.

Examining High Magnitude Grass Pollen Episodes at Worcester, United Kingdom, Using Back-Trajectory Analysis

Trajectory analysis is a valuable tool that has been used before in aerobiological studies, to investigate the movement of airborne pollen. This study has employed back-trajectories to examine the four highest grass pollen episodes at Worcester, during the 2001 grass pollen season. The results have shown that the highest grass pollen counts of the 2001 season were reached when air masses arrived from a westerly direction. Back-trajectory analysis has a limited value to forecasters because the method is retrospective and cannot be employed directly for forecasting. However, when used in conjunction with meteorological data this technique can be used to examine high magnitude events in order to identify conditions that lead to high pollen counts.

Characteristics of Grass Pollen Seasons in Areas of Southern Spain and the United Kingdom

Spatial and temporal variations in daily grass pollen counts and weather variables are described for two regions with different bio-geographical and climatic regimes, southern Spain and the United Kingdom. Daily average grass pollen counts are considered from six pollen-monitoring sites, three in southern Spain (Ciudad Real, Córdoba and Priego) and three in the United Kingdom (Edinburgh, Worcester and Cambridge). Analysis shows that rainfall and maximum temperatures are important factors controlling the magnitude of the grass pollen season in both southern Spain and the United Kingdom, and that the strength and direction of the influence exerted by these variables varies with geographical location and time.

Temporal and Georaphical Variations in Grass Pollen Seasons in Areas of Western Europe: an Analysis of Season dates at Sites of the European Pollen Information System.

Geographical and temporal variations in the start dates of grass pollen seasons are described for selected sites of the European Pollen Information Service. Daily average grass pollen counts are derived from Network sites in Finland, the Netherlands, Denmark, United Kingdom, Austria, Italy and Spain, giving a broad longitudinal transect over Western Europe. The study is part of a larger project that also examines annual and regional variations in the severity, timing of the peak and duration of the grass pollen seasons. For several sites, data are available for over twenty years enabling long term trends to be discerned. The analyses show notable contrasts in the progression of the seasons annually with differing lag times occurring between southern and northern sites in various years depending on the weather conditions. The patterns identified provide some insight into geographical differences and temporal trends in the incidence of pollinosis. The paper discusses the main difficulties involved in this type of analysis and notes possibilities for using data from the European Pollen Information service to construct pan European predictive models for pollen seasons.