Non-culturable Rhizobia and diverse endophytic bacteria co-inhabiting wild legume nodules.

Analyses of the bacterial content of legume nodules does generally yield cultures of the symbiotic partner responsible of nodule induction, invasion and nitrogen fixation. Species involved in such interaction include examples among the alpha and beta-rhizobia. In the present survey, exploring the nodular content of several species of wild legumes in the Mediterranean basin, we witnessed a consistent occurrence of the following situation: the bacteria which could be isolated from surface- sterilized nodules do not represent the actual nodule-inducing microorganism but rather a variety of endophytic invaders. Their presence in the nodules appeared related to their more general and systemic distribution throughout the whole plant, in close analogy with the situation observed in many other herbaceous and woody species which host an array of on-pathogenic endophytic taxa. At the same time the presence and quantitative abundance of the real nodule symbiont in the wild legumes covered by our study could be demonstrated by 16S rRNA gene PCR and nucleotide sequencing, which revealed that, despite their failure to grow on plates, alpha-proteobacterial rhizobia do represent the bona-fide symbiont responsible of nodule organogenesis.