Diversity and distribution of Frankia strains symbiotic with Ceanothus in California

Abstract

Frankia strains symbiotic with Ceanothus present an interesting opportunity to study the patterns and causes of Frankia diversity and distribution within a particular host infectivity group. We intensively sampled Frankia from nodules on Ceanothus plants along an elevational gradient in the southern Sierra Nevada of California, and we also collected nodules from a wider host taxonomic and geographic range throughout California. The two sampling scales comprised 36 samples from eight species of Ceanothus representing six of the seven major biogeographic regions in and around California. The primary objective of this study was to use a quantitative model to test the relative importance of geographic separation, host specificity, and environment in influencing the identity of Ceanothus Frankia symbionts as determined by ribosomal DNA sequence data. At both sampling scales, Frankia strains symbiotic with Ceanothus exhibited a high degree of genetic similarity. Frankia strains symbiotic with Chamaebatia (Rosaceae) were within the same clade as several Ceanothus symbionts. Results from a classification and regression tree model used to quantitatively explain Frankia phylogenetic groupings demonstrated that the only significant variable in distinguishing between phylogenetic groups at the more local sampling scale was host species. At the regional scale, Frankia phylogenetic groupings were explained by host species and the biogeographic province of sample collection. We did not find any significant correspondence between Frankia and Ceanothus phylogenies indicative of coevolution, but we concluded that the identity of Frankia strains inhabiting Ceanothus nodules may involve interactions between host species specificity and geographic isolation.

FIG. 1.

Geographic locations, elevations, and host plants from which nodules were collected. The graph on the right summarizes host species and elevations from the intensive regional sampling of Ceanothus nodules, while the graph on the left summarizes the sampling of three host genera across California. Species names are listed in Table 2. Samples represent five of the six major geographic regions of the California Floristic Province and one region of the Great Basin Floristic Province (per reference 14).

FIG. 2.

Neighbor-joining tree based on IGS and 23S sequence data from Frankia strains symbiotic with Ceanothus collected from the intensive regional sampling in the southern Sierra Nevada. The tree is rooted with a sequence from the Alnus symbiont strain AcN14a (21). Bootstrap values represent 1,000 replicates; values are shown as percentages only when they exceed 50%. Branch lengths correspond to sequence differences as indicated by the scale bar. Designations at terminal nodes consist of a four-letter host species code (CEIN, C. integerrimus; CEPA, C. parvifolius; CECO, C. cordulatus; CECU, C. cuneatus; CELE, C. leucodermis) followed by the elevation (in meters) where the plant was growing. Details of sample origins are presented in Table 2. *, nodules formed in a greenhouse in Seattle on C. cordulatus seedlings grown in field-collected soil as described in the text. ^1,2, sympatric paired samples.

FIG. 3.

Neighbor-joining tree based on IGS and 23S sequence data from Frankia strains symbiotic with Ceanothus, Alnus, and Chamaeba tia collected from six major biogeographic regions throughout California. Sequence from Streptomyces griseus was used to root the tree. Bootstrap values represent 1,000 replicates; only values above 50% are shown. Branch lengths correspond to sequence differences as indicated by the scale bar. Designations at terminal nodes consist of a four-letter host species code followed by the elevation (in meters) where the plant was growing. Plant groups are based on a molecular phylogeny of Ceanothus (17), with groupings inferred on the basis of morphological characters for C. greggi (CEGR), C. leucodermis, and C. spinosus (CESP). Frankia phylogenetic groups were assigned for the purposes of statistical analysis, and so groups with a membership of <3 were not created even when supported by bootstrapping. ^1,2,3, sympatric paired samples. CETH, C. thyrsifolius; CHFO, Chamaebatia foliolosa; ALRH, Alnus rhombifolia; ALVI, Alnus viridis; ALIN, Alnus incana; ALRU, A. rubra. Other host species codes are as explained in the legend to Fig. 2.

FIG. 4.

Classification tree of relative importance of variables used to explain the phylogenetic groupings of Frankia strains associated with Ceanothus and Chamaebatia shown in Fig. 3. Values at terminal nodes are the phylogenetic groups from Fig. 3. Of the 12 independent variables used in the analysis (host species, elevation, geographic region, geographic subregion, soil pH, latitude, plant phylogenetic grouping, and five predicted climate parameters for each sample location as described in the text), only host species and geographic region were significant.

FIG. 5.

Neighbor-joining tree based on IGS sequence data comparing previously published sequences (19, 32) with those for a subset of Frankia strains analyzed in this study that were chosen to be representative of the geographic regions covered. Previously published sequences (19, 32) are marked by asterisks. The tree is rooted with a sequence from Streptomyces griseus. Bootstrap values represent 1,000 replicates; only values above 50% are shown. Branch lengths correspond to sequence differences as indicated by the scale bar. Designations at terminal nodes consist of a four-letter host species code followed by the elevation (in meters) where the plant was growing. Plant groups are based on a molecular phylogeny of Ceanothus (17), with groupings inferred on the basis of morphological characters for C. greggi, C. leucodermis, C. parvifolius, and C. spinosus (CESP). Identical superior numbers indicate sympatric paired samples. CEVE, C. velutinus; CEPU, C. pumilus; CEPR, C. prostratus; CETH, C. thyrsifolius. Other host species codes are as explained in the legends to Fig. 2 and 3.