# Phylosymbiosis in the Rhizosphere Microbiome Extends to Nitrogen Cycle Functional Potential
## 3 Results
### Microbiome Composition  
Change in rhizosphere microbiome composition, after rarefaction, correlated with the plant phylogeny. First, as expected, within pots, microbiomes displayed more similar taxonomic composition (at the genus level) than across pots ([PERSON]'s \(t\)-test, \(p<0.001\)) (Figure 2A and Supplementary Figure S1). Comparisons between samples from the same pot were removed from all subsequent analysis. Next, we focused on the microbiomes of plants from the same species (i.e., two collections of _Leptochloa dubia_) and the same genus (i.e., two _Distichlis_ species and three _Sporobolus_ species) We found that microbiomes from different pots but from closely related plants had more similar microbiomes than pairs of randomly selected microbiomes. Specifically, pairwise comparisons between samples from different pots revealed that variation in the microbiome composition was in the range of variation observed between same-pot replicates, except for one _Sporobolus_ sample (Figure 2A). However, when we considered all the pairwise comparisons, after excluding the comparison among replicates, the dissimilarity between pairs of rhizosphere microbiomes increased with the phylogenetic distance among the plant hosts (r\({}_{\text{Pearson}}\) = 0.2, \(p<0.001\); r\({}_{\text{Spearman}}\) = 0.22, \(p<0.001\), Figure 2A). This overall weak correlation suggests that a large fraction of the microbiome is either conserved across samples, as expected in a common garden experiment, or does not mirror the plant phylogeny. Finally, we used a linear regression (MicroComp \(\sim\)PlantPhylo) to assess the linkage between the variation in the microbial community composition and the phylogenetic distance among pairs of plants (slope = 0.658, \(p<0.001\), std.error = 0.063; Figure 2A). Together, these results show that more closely related plants have more similar rhizosphere microbial communities. However, the microbial communities from replicated soil samples from the same pot and samples from the same species/genus, although highly similar, were not identical (Figure 2A). We also investigated the effect of sample geography based on where the plant was originally collected but found no significant difference when investigating pairwise comparisons of the microbiome from plants derived from the same continent and plants from different continents (Supplementary Figure S2A). Across samples, a total of 1,276,324,785 sequences (post QC) were analyzed (Supplementary Table S2) and the rhizosphere microbiome was dominated by Proteobacteria, with annotated sequences ranging from 51.2 to 69.6%. This was also reflected at the genus level, as the five most abundant genera, _Bradyrhizobium_, _Rhodpseudomonas_, _Burkholderia_, _Mesorhizobium_, and _Optituts_ (phylum Verrucomicrobia), accounted for an  
Figure 2: Relationship between phylogenetic distance among pairs of plants and the microbial community dissimilarity (**A**) and variation in the functional potential for N cycling in the corresponding microbial communities (**B**). Comparisons among technical replicates (same plant, black circles), samples from the same species (triangles), and from the same genus (squares) are highlighted. average of 15.56% of the annotated reads (Supplementary Figure S1). After rarefaction of the taxonomically identified sequences (\(n\) = 3,127,337 sequences/sample), 481 bacterial genera were identified. Although some abundant lineages were identified, most genera were associated with a low number of sequences. Globally, the Shannon diversity index (\(H\)), computed at the genus level, ranged from 4.95 to 5.40 (Supplementary Figure S1A). Across samples, most genera accounting for >0.5% of the reads displayed low variation in their frequency (Supplementary Figure S1).