What’s under the Christmas Tree? A Soil Sulfur Amendment Lowers Soil pH and Alters Fir Tree Rhizosphere Bacterial and Eukaryotic Communities, Their Interactions, and Functional Traits

In this study, we describe the legacy effects of a soil sulfur amendment experiment performed 6 years prior and the resulting alterations to the rhizosphere communities of fir trees on a Christmas tree plantation. The pH of bulk soil was ∼1.4 pH units lower than that of untreated soils and was associated with reduced Ca, Mg, and organic matter contents. Similarly, root chemistry differed due to the treatment, with roots in sulfur-amended soils showing significantly higher Al, Mn, and Zn contents and reduced levels of B and Ca. 16S rRNA and 18S rRNA gene sequencing was pursued to characterize the bacterial/archaeal and eukaryotic communities in the rhizosphere soils. The treatment induced dramatic and significant changes in the microbial populations, with thousands of 16S rRNA gene sequence variants and hundreds of 18S rRNA gene variants being significantly different in relative abundances between the treatments. Additionally, co-occurrence networks showed that bacterial and eukaryotic interactions, network topology, and hub taxa were significantly different when constructed from the control and treated soil 16S and 18S rRNA gene amplicon libraries. Metagenome sequencing identified several genes related to transport proteins that differentiated the functional potentials of the communities between treatments, pointing to physiological adaptations in the microbial communities for living at altered pH. These data show that a legacy of soil acidification increased the heterogeneity of the soil communities as well as decreasing taxon connections, pointing to a state of ecosystem instability that has potentially persisted for 6 years. IMPORTANCE We used sulfur incorporation to investigate the legacy effects of lowered soil pH on the bacterial and eukaryotic populations in the rhizosphere of Christmas trees. Acidification of the soils drove alterations of fir tree root chemistry and large shifts in the taxonomic and functional compositions of the communities. These data demonstrate that soil pH influences are manifest across all organisms inhabiting the soil, from the host plant to the microorganisms inhabiting the rhizosphere soils. Thus, this study highlights the long-lasting influence of altering soil pH on soil and plant health as well as the status of the microbiome.