Genetic composition of symbiotic and free-living Frankia
communities in Alnus–Frankia networks
We obtained a total of 352 ASVs from 138 samples consisting of riparian
soil, rhizosphere soil, and root nodules (Fig. S1). Of the total ASVs,
90.34 % were unique singleton variants, which occurred once in 138
samples. We found profoundly diverse ASVs from rhizosphere and riparian
soils (rhizosphere: 228 ASVs, n = 54; riparian: 103 ASVs,n = 12). In contrast, in root nodules, the total number of ASVs
was quite limited (38 ASVs, n = 72). These results suggest that
host individuals establish symbiosis only with limited Frankiagenotypes in soil.
To classify the Frankia genetic variants, we used OTU methods
based on the genetic similarity of the nif D-K loci. A total of 19
OTUs were obtained at a 97% similarity threshold. Consequently, most
ASVs from root nodules and riparian soils converged to three OTUs
(OTU01, 02, and 03), while a variety of OTUs were found in the
rhizosphere soil (Fig. 2). Consistently, the sample-based rarefaction
curves showed that the number of Frankia OTUs did not completely
reach saturation but were significantly greater in number in
rhizospheres than in root nodules after standardization of the sample
size (Fig. S3). In addition, the number of Frankia OTUs was lower
in riparian soils than in rhizosphere soils (Fig. S3). These findings
suggest that the occurrence of host plants promote the local assembly of
nitrogen-fixing bacterial symbionts in soil rather than homogeneity
dispersion of diverse bacterial strains in a natural forest.
Moreover, the phylogenetic trees indicated that the Frankia OTUs
detected in our study covered a wide variety of Alnus -infecting
strains (Fig. S2). The most abundant strains (i.e., OTU01, 02, and 03)
belonged to phylogenetically distant clades (Fig. S2), indicating that
the Alnus host population accepts genetically diverse bacterial
symbionts.