South African Papilionoid Legumes Are Nodulated by Diverse Burkholderia with Unique Nodulation and Nitrogen-Fixation Loci

Abstract

The root-nodule bacteria of legumes endemic to the Cape Floristic Region are largely understudied, even though recent reports suggest the occurrence of nodulating Burkholderia species unique to the region. In this study, we considered the diversity and evolution of nodulating Burkholderia associated with the endemic papilionoid tribes Hypocalypteae and Podalyrieae. We identified distinct groups from verified rhizobial isolates by phylogenetic analyses of the 16S rRNA and recA housekeeping gene regions. In order to gain insight into the evolution of the nodulation and diazotrophy of these rhizobia we analysed the genes encoding NifH and NodA. The majority of these 69 isolates appeared to be unique, potentially representing novel species. Evidence of horizontal gene transfer determining the symbiotic ability of these Cape Floristic Region isolates indicate evolutionary origins distinct from those of nodulating Burkholderia from elsewhere in the world. Overall, our findings suggest that Burkholderia species associated with fynbos legumes are highly diverse and their symbiotic abilities have unique ancestries. It is therefore possible that the evolution of these bacteria is closely linked to the diversification and establishment of legumes characteristic of the Cape Floristic Region.

Biogeographical Patterns of Legume-Nodulating Burkholderia spp.: from African Fynbos to Continental Scales

“…While the global Burkholderia diversity was structured for the nodulation genes at the legume subfamily level, an interaction of rhizobia, host legumes, and geographic distribution was not shown at a regional scale, showing widely spread and locally diverse Burkholderia populations in the fynbos. Our results corroborate a previous study demonstrating the widespread occurrence of Burkholderia and the absence of a site sampling effect on the rhizobial diversity of selected Hypocalypteae and Podalyrieae species (27,32). Using geographic distances as a proxy for population connectivity, genetic variation is expected to correlate positively with the sampling site distances.…”

“…For both chromosomal and nodulation genes, with the latter symbiotic genes determining host specificity (68), a relaxed association among genetically similar rhizobia and different legume species, genera, and tribes was demonstrated. The variation of host-Burkholderia interactions corroborates a previous rhizobial screening in selected legume genera of the tribes Hypocalypteae (Hypocalyptus) and Podalyrieae (Cyclopia, Podalyria, Virgilia) (27,32). In South America, a similar relaxed host-specific interaction has been described for Burkholderia bacteria and their mimosoid hosts (10,62).…”

“…In the tribe Podalyrieae, a strong preference for Burkholderia is observed, showing all legume species and genera (except for Calpurnia, which is not endemic to the fynbos; Table 2; see Fig. S2 in the supplemental material) strictly nodulated with Burkholderia (12,26,27). Other common plant groups, such as the tribes Crotalarieae and Indigofereae, also contain Burkholderia-philous species, although (closely related) legume lineages within the same tribes and co-occurring in the similar habitats have been recorded with classical alpha-rhizobial lineages (26 and references therein).…”

“…In our inoculation experiment, siratro and P. pinnata nodulated in soils collected from South Africa and Malawi, whereas P. calyptrata, I. filifolia, and D. lignosus were nodule free in Malawian soil. The inability to form nodules in Malawian soil suggests that these legumes, known to exhibit a strong host preference for Burkholderia (26,27,83), did not find their specific Burkholderia symbionts in the Malawian (savanna) soil, which had a substantially higher pH than the Cape soil. The occurrence and success of Burkholderia in South African (fynbos) soils, but also in the South American Cerrado/Caatinga biomes, can be linked with the general ecological adaptation of these symbionts to acidic soil conditions, which may play a prominent role as an ecological driver of rhizobial diversity (21,29,30,35).…”

“…In South Africa, Burkholderia symbionts are widespread and associated with diverse lineages of the tribes Crotalarieae (23)(24)(25)(26), Hypocalypteae (27,28), Indigofereae (26), Phaseoleae (26,29,30), and Podalyrieae (26,27,31,32), indicating that the South African soils are an important reservoir for nodulating Burkholderia, and thus this needs to be explored further for new candidate species. With the exception of Burkholderia phymatum strains nodulating the nonnative crop species Phaseolus vulgaris (common bean) in Moroccan soils (33), the legume-Burkholderia symbiosis in Africa has only been reported in a range of sites within the fynbos region, supporting the idea of the Cape region as an exclusive biodiversity hot spot for the symbiosis (12).…”