Diversity of Frankia Populations in Root Nodules of Sympatrically Grown Alnus Species
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The potential role played by the host plant or the soil type in the selection of symbiotic, nitrogen-fixing Frankia strains from soil for root nodule formation was assessed by using molecular techniques. The diversity of Frankia populations in the root nodules of twelve Alnus species grown sympatrically was analyzed using rep-PCR (Repetitive Extragenomic Palindromic polymerase chain reaction). Analysis of 120 root nodules, i.e. 10 root nodules from each plant species revealed three rep-profiles which indicated the presence of only three Frankia strains in the root nodules. One profile (further referred to as Type I) was most abundant and was identical within and among the nodules of nine species (A. incana, A. japonica, A. glutinosa, A. tenuifolia, A. rugosa, A. rhombifolia, A. mandsurica, A. maritima and A. serrulata). Type II profiles were found in all nodules of two plant species (A. hirsuta and A. glutinosa pyramidalis), whereas the type III profile was unique for Frankia populations in nodules of A. rugosa americana. No variation was detected in Frankia populations among the root nodules of a single plant species, i.e., all nodules from one plant species had identical rep-profiles. Comparative sequence analyses of nifH gene fragments from three Frankia strains representing each rep-profile (Type I: A. tenuifolia; Type II: A. glutinosa pyramidalis; Type III: A. rugosa americana) revealed differences, with Type I and II clustering with Frankia strains of subgroup AII (represented by strain Ag45/Mut15) and Type III with Frankia of subgroup AI (represented by strain ArI3). Comparative sequence analysis with 73 clones from a nifH gene clone library from frankiae in soil where these 12 alder species were growing, however, revealed that none of these sequences represented frankiae detectable in soil. Sequences from all 73 clones clustered in subgroup AI with similarity values of 93, 97, and 99.6 %, to sequences from populations in nodules of A. tenuifolia, A. glutinosa pyramidalis and A. rugosa americana, respectively. Additional analyses of nodule populations from alders growing on different soils demonstrated the presence of different Frankia populations in nodules for each soil, with populations showing identical sequences in nodules from the same soil, but differences between plant species. These results suggest that soil environmental conditions and host plant species have a role in the selection of Frankia strains for root nodule formation and this selection is not a function of the abundance of a Frankia strain in soil.