Diversity of bradyrhizobial T3SS systems and their roles in symbiosis with peanut (Arachis hypogaea) and Vigna species (V. radiata and V. mungo)

Abstract

Symbiosis between Bradyrhizobium strains isolated from Lao People’s Democratic Republic (Lao PDR) and intercropped legumes (Arachis hypogaea, Vigna radiata, and V. mungo) was regulated by the type III secretion system (T3SS), which delivers effector proteins (T3Es) into host plant cells to modulate nodulation. To explore this mechanism, we sequenced and analyzed seven Bradyrhizobium genomes, identifying putative T3Es across five T3SS groups (G.1–G.5), which were classified based on the sequence of rhcN, a conserved ATPase gene essential for T3SS function. Phylogenetic analysis of rhcN more closely reflected the evolutionary relationships of nodulation genes than those based on 16S rRNA or whole-genome comparisons, underscoring its symbiotic relevance. Functional assays using rhcN mutants revealed group-specific effects on nodulation; G.1 strains showed neutral effects on A. hypogaea, negative effects on V. radiata, and positive effects on V. mungo. G.2 strains consistently promoted nodulation across all hosts and lacked effectors related to SUMO (small ubiquitin-like modifier) pathways, which have been implicated in host defense regulation. G.3 strains reduced nodulation in A. hypogaea but enhanced it in Vigna species. G.4 strains suppressed nodulation in A. hypogaea, and G.5 strains inhibited nodulation across all tested legumes. These findings highlight the diversity in T3SS organization, effector composition, and symbiotic responses among native Bradyrhizobium strains. The identification of known and uncharacterized effectors suggests roles in host compatibility and specificity. These strains, along with their effector profiles, provide a foundation for future functional studies to better understand T3SS-mediated interactions and support the development of targeted inoculants for legume hosts.