Soil Frankia nifH analyses. Soil Frankia

Actinorhizal plants can form nitrogen-fixing root nodules in symbiosis with members of the soil-dwelling actinobacterial genus Frankia, with members of specific Frankia taxonomic clusters nodulating plants in corresponding host infection groups. Frankia diversity was assessed in root nodules of different host plant species and related to the abundance and relative distribution of indigenous frankiae in rhizosphere soils. Differential nodulation was observed for plant species within host infection groups, with some host species being nodulated (Alnus glutinosa, A. cordata, Shepherdia argentea, Casuarina equisetifolia) while others were not (A. viridis, Hippophaë rhamnoides). Sequence analyses of nifH gene fragments retrieved from nodule populations revealed a total of eight different sequences, two of which characterized frankiae in nodules from S. argentea, three sequences characterized frankiae in nodules from A. glutinosa, and five sequences characterized frankiae in nodules from A. cordata of which two were shared with A. glutinosa. Quantitative PCR (qPCR) of soil DNA demonstrated that vegetation generally increased abundance of frankiae in soil, with identical results within treatments independent of the target gene (i.e. the nifH or the 23S rRNA gene). The soil analyses found frankiae of clusters 1b, a newly assigned cluster 1e, and cluster 3 only, while clusters 1a/d, 1c, 2 and 4 were not detected. Targeted Illumina sequencing of Frankia-specific nifH gene fragments retrieved a total of 24 unique sequences from all rhizosphere soils and nodules, with seven of these sequences representing generally more than 90% of the reads obtained in the majority of samples. Only two of the seven sequences were identical to those found in nodules, one representing cluster 3 frankiae (generally 3 to 5 % of the reads) and the second representing cluster 1e frankiae (generally 4 to 9% of the reads). The two most abundant sequences that represented cluster 3 frankiae were not found in root nodules. Abundance in soils did not correlate to presence in nodules. These results demonstrate large differences between detectable Frankia populations in soil and those in root nodules suggesting that root nodule formation is not a function of abundance of specific Frankia populations in soils.