Quantitative DNA Analyses for Airborne Birch Pollen.

Isabell Müller-Germann,Heike Vogel,Ulrich Pöschl,Bernhard Vogel,Viviane R Després,Janine Fröhlich-Nowoisky,Andreas Pauling,Maosheng Yao

doi:10.1371/journal.pone.0140949

Abstract

Birch trees produce large amounts of highly allergenic pollen grains that are distributed by wind and impact human health by causing seasonal hay fever, pollen-related asthma, and other allergic diseases. Traditionally, pollen forecasts are based on conventional microscopic counting techniques that are labor-intensive and limited in the reliable identification of species. Molecular biological techniques provide an alternative approach that is less labor-intensive and enables identification of any species by its genetic fingerprint. A particularly promising method is quantitative Real-Time polymerase chain reaction (qPCR), which can be used to determine the number of DNA copies and thus pollen grains in air filter samples. During the birch pollination season in 2010 in Mainz, Germany, we collected air filter samples of fine (<3 μm) and coarse air particulate matter. These were analyzed by qPCR using two different primer pairs: one for a single-copy gene (BP8) and the other for a multi-copy gene (ITS). The BP8 gene was better suitable for reliable qPCR results, and the qPCR results obtained for coarse particulate matter were well correlated with the birch pollen forecasting results of the regional air quality model COSMO-ART. As expected due to the size of birch pollen grains (~23 μm), the concentration of DNA in fine particulate matter was lower than in the coarse particle fraction. For the ITS region the factor was 64, while for the single-copy gene BP8 only 51. The possible presence of so-called sub-pollen particles in the fine particle fraction is, however, interesting even in low concentrations. These particles are known to be highly allergenic, reach deep into airways and cause often severe health problems. In conclusion, the results of this exploratory study open up the possibility of predicting and quantifying the pollen concentration in the atmosphere more precisely in the future.

Highlights

Pollen allergies affect up to 30% of the human population in industrialized countries [1] and are today the most common form of seasonal respiratory diseases in Europe [2,3]
While the DNA on all coarse and fine particle filter samples could be quantified with the internal transcribed spacer (ITS) primer pair, only ~90% of the coarse and ~45% of the fine particle filters were quantifiable with the BP8 primer pair
In comparison to Betula pendula the occurrence is very small, and both species can be amplified by the ITS primer pair, likely negligible

Summary

Introduction

Pollen allergies affect up to 30% of the human population in industrialized countries [1] and are today the most common form of seasonal respiratory diseases in Europe [2,3]. The establishment and enhancement of accurate pollen quantification and forecasting systems are essential for the improvement of public health. Pollen forecasts are mainly based on the results provided by microscopic analyses of pollen traps, in which pollen and spores are collected [5] and identified based on optical recognition of specific morphological features [6]. Pollen forecasts are based on weather forecasts and the described pollen identification data, which often only comes with a time delay of one or two days. This method is still the standard for national pollen networks covering most of the European continent [1,5]

Methods

Results

Conclusion