in phytochemical components and environmental factors, resulting in alteration in the phyllosphere bacterial neighborhood; as a result, specific bacteria occurred in stands of specific ages. A LDA of impact size (LEfSe) evaluation was performed to detect the bacterial taxa that showed important variations in relative abundance (BRPF2 Inhibitor Purity & Documentation Figures 4A,B and Supplementary Figure 4). Members of the genus Massilia were rare in sapling stands (SM5) but showed higher relative abundance in mature stands (SM25) (p 0.001) (Figure 4C). Pantoea species had been uncommon in overmatureFrontiers in Plant Science | frontiersin.orgSeptember 2021 | Volume 12 | ArticleSun et al.Phyllosphere Bacterial Communities and MetabolomesFIGURE two | Beta DP Inhibitor supplier diversity in the phyllosphere bacterial community of Chinese fir. (A) Scatterplot of axes 1 and 2 from a PCoA determined by unweighted UniFrac dissimilarity matrix; (B) unweighted ANOSIM; (C) UPGMA dendrogram. SM5, SM15, SM25, and SM35 represent stand ages of five, 15, 25, and 35 years, respectively.stands (SM35) and showed the highest relative abundance in SM25 (p 0.001) (Figure 4D). Members of the family members Burkholderiaceae (p 0.001), the order Betaproteobacteriales (p 0.001) and the class Gammaproteobacteria (p 0.001) showed the lowest relative abundances in SM5 and SM35, and the highest relative abundances in SM25 (Figures 4E ). Species on the loved ones Rhodanobacteraceae (p 0.001) as well as the genus Luteibacter (p 0.001) were detected in the younger stands (SM5 and SM15) but were barely detected in the older stands (SM25 and SM35) (Supplementary Figures 3A,B). Members of the phylum Actinobacteria have been detected in all 4 stands but showed decrease relative abundance in SM25 (p 0.001) (Supplementary Figure 3C). Some members ofthe phylum Acidobacteria, which had been also detected in all 4 stands, showed reduced relative abundance in SM5 (p 0.001) (Supplementary Figure 3D).Predicted Functions of Bacterial CommunitiesThe application PICRUSt was used to predict variation in functional genes in the phyllosphere bacterial communities. The bacterial communities involved inside the metabolic pathways for immune illnesses, and worldwide and overview maps showed a considerable difference between SM5 and SM15 (p 0.05; Figure 5A). Similarly, bacterial communities showed important differences in glycan biosynthesis and metabolism, and metabolism of terpenoids and polyketides between SM15 and SM25 (p 0.05;Frontiers in Plant Science | frontiersin.orgSeptember 2021 | Volume 12 | ArticleSun et al.Phyllosphere Bacterial Communities and MetabolomesFIGURE three | Abundance of phyllosphere bacterial communities with relative abundance 0.8 as classified to (A) family members; (B) genus; and (C) species taxonomic levels. (D) The top 30 bacterial orders identified by the predictive Random Forest model using the imply decreasing Gini score. SM5, SM15, SM25, and SM35 represent stand ages of five, 15, 25, and 35 years, respectively.Figure 5B). With an increase in stand age from 25 to 35 years, adjustments inside the phyllosphere bacterial community impacted additional metabolic pathways. For 12 metabolic pathways, the bacterial neighborhood showed important differences between SM25 and SM35 (p 0.05; Figure 5C). Significant variations in bacterial communities involved in extra metabolic pathways were detected for other pairwise comparisons of stands (Supplementary Figure four). Detailed information around the predicted functional genes inside the phyllosphere communities, particularly thirdtier KEGG pathways, i