Pollen-associated microbiome correlates with pollution parameters and the allergenicity of pollen

Pollen allergies have been rapidly increasing over the last decades. Many allergenic proteins and non-allergenic adjuvant compounds of pollen are involved in the plant defense against environmental or microbial stress. The first aim of this study was to analyze and compare the colonizing microbes on allergenic pollen. The second aim was to investigate detectable correlations between pollen microbiota and parameters of air pollution or pollen allergenicity. To reach these aims, bacterial and fungal DNA was isolated from pollen samples of timothy grass (Phleum pratense, n = 20) and birch trees (Betula pendula, n = 55). With this isolated DNA, a terminal restriction fragment length polymorphism analysis was performed. One result was that the microbial diversity on birch tree and timothy grass pollen samples (Shannon/Simpson diversity indices) was partly significantly correlated to allergenicity parameters (Bet v 1/Phl p 5, pollen-associated lipid mediators). Furthermore, the microbial diversity on birch pollen samples was correlated to on-site air pollution (nitrogen dioxide (NO2), ammonia (NH3), and ozone (O3)). What is more, a significant negative correlation was observed between the microbial diversity on birch pollen and the measured NO2 concentrations on the corresponding trees. Our results showed that the microbial composition of pollen was correlated to environmental exposure parameters alongside with a differential expression of allergen and pollen-associated lipid mediators. This might translate into altered allergenicity of pollen due to environmental and microbial stress

Comparison of fungal pattern on pollen from birch and timothy grass.

<p><b>A)</b> PCoA (Bray-Curtis-Index) of fungal pattern on birch pollen (<i>Betula pendula</i>; n = 31; black) and timothy grass pollen (<i>Phleum pratense</i>; n = 16; grey) show hardly overlapping 95%-confidence. The significant differences were confirmed by ANOSIM (R = 0.51, p = 0.0001). <b>B)</b> The cluster dendrogramm (Bray-Curtis-Index) demonstrates the separation of the two pollen species due to fungal pattern. <b>C,D)</b> Boxplots of indices weighing species due to their extent of abundance in fungal community on birch- and timothy grass-pollen. In comparison the two groups differed significantly due to low abundant species (Shannon-index, p = 0.006), and also due to high abundant species of fungal composition (Simpson-index, p = 0.005).</p

Spearman correlations of bacterial diversity on birch pollen to allergenicity parameter.

<p>Scatterplot including linear trend line, spearman correlation values rho and significance level p of diversity indices (Shannon H/Simpson 1-D) and parameter (number of fragments n(tRFs)) of bacterial pattern on birch pollen (<i>Betula pendula</i> 2014, n = 55) with allergen concentration in pollen. <b>A-D)</b> Correlation of n(tRFs) and allergenicity parameters Bet v 1 (r_s = 0.28, p = 0.038), PALM_LTB4 (r_s = 0.14, p = 0.33), PALM_PGE2 (r_s = 0.17, p = 0.22) and NADPH oxidase (r_s = -0.04, p = 0.79). <b>E-H)</b> Correlation of Simpson diversity Index 1-D and allergenicity parameters Bet v 1 (r_s = 0.25, p = 0.068), PALM_LTB4 (r_s = 0.08, p = 0.53), PALM_PGE2 (r_s = .12, p = 0.366) and NADPH oxidase (r_s = 0.2, p = 0.19). <b>I-L)</b> Correlation of Shannon diversity Index H and allergenicity parameters Bet v 1 (r_s = 0.14, p = 0.317), PALM_LTB4 (r_s = -0.36, p = 0.006), PALM_PGE2 (r_s = -0.04, p = 0.0015) and NADPH oxidase (r_s = 0.2, p = 0.19).</p

Comparison of bacterial and fungal pattern on pollen collected in rural and urban sample sites.

<p>PCoA performed to compare the microbial composition on pollen obtained from rural (grey) and urban areas (black). <b>A)</b> Bacterial composition on birch pollen. <b>B)</b> Bacterial composition on timothy grass pollen. <b>C)</b> Fungal composition on birch pollen. <b>D)</b> Fungal composition on timothy grass pollen.</p

Comparison of bacterial pattern on pollen from birch and timothy grass.

<p><b>A)</b> PCoA (Bray-Curtis-Index) of bacterial pattern on birch pollen (<i>Betula pendula</i>; n = 55; black) and timothy grass pollen (<i>Phleum pratense</i>; n = 20; grey) show hardly overlapping 95%-confidence intervals. The significant differences were confirmed by ANOSIM (R = 0.81, p = 0.0001). <b>B)</b> In the cluster dendrogramm (Bray-Curtis) pollen from both species are forming separated groups also attesting the significant difference. <b>C,D)</b> Boxplots of indices weighing species due to their extent of abundance in bacterial community on birch- (<i>Betula pendula</i>; n = 55) and timothy grass-pollen (<i>Phleum pratense</i>, n = 20). In comparison the two groups differs significantly due to low abundant species (Shannon-index, p = 0.009), but not significantly due to high abundant species of bacterial composition (Simpson-index, p = 0.78).</p