Azoarcus spp. and their interactions with grass roots

The current knowledge on the divergence within the genus Azoarcus and about interactions with grasses is summarized. Grass-associated members of this genus of diazotrophs have only been isolated from a salt- and flood-tolerant pioneer plant in Pakistan, Kallar grass (Leptochloa fusca (L.) Kunth). Members of these bacteria belong to the beta subclass of the Proteobacteria, most closely related to purple bacteria such as Rhodocyclus purpureus. The isolates from one single host plant showed a surprising divergence, consisting of five groups of Azoarcus distinct at the species level. Molecular diagnostic tests, which are based on 16S ribosomal RNA sequences, allowed preliminary assignment of isolates to Azoarcus by PCR amplification and sequencing of PCR products. Moreover, the moleculer tests enabled us to detect an unculturable strain in Kallar grass roots, stressing that classical cultivation techniques at times fail to detect some groups of the microbial population. Using similar techniques, sequences rooting in the Azoarcus clade were also detected in field-grown rice, indicating that the natural host range might extend to rice. In gnotobiotic laboratory cultures, a member of Azoarcus is able to colonize rice roots endophytically: bacteria invade the roots in the zone of elongation and differentiation, colonize the cortex intra- and inter-cellularly, and penetrate deeply into the vascular system, entering xylem vessels, allowing systemic spreading into the rice shoot. Recently, we detected expression of nitrogenase of Azoarcus cells inside roots of rice seedlings, a result encouraging us to analyze interactions with rice in detail.