The spontaneous nodulation of alfalfa

The establishment of symbiosis between leguminous plants and the soil bacteria Rhizobia is investigated using the Nar or spontaneous nodulation phenomenon, i.e. ability of some plant to develop nodulations in the absence of Rhizobium. It is shown that he spontaneous nodule organogenesis is initialized in both Nar and non-Nar plants but more or less blocked further (at the nodule primordia stage), suggesting a regulation of the nodule morphogenesis after the induction of the primordium. Purified nodulation factors were shown to be implied in mitogenic phenomenon, increasing the nodulation phenomenon. Finally, the comparison between spontaneous and Rhk-obium-induced nodulation suggests that the cortical divisions do not result from a bacterial activation but rather from a spontaneous reactivity of the cortex. words: Medicago sativa, alflafa, spontaneous nodulation, NAR, Rhizobium Interaction between leguminous plants and the soil bacteria Rhizobia results in the development of a newly formed plant organ, the root nodule, in which bacteria reduce atmospheric nitrogen into ammonia. The nodular organogenesis has been extensively studied by using legume and bacterial mutants, blocked at various steps of this developmental program, or by adding purified molecules, such as bacterial nodulation factors or phytohormones. Another useful tool arose a few years ago, with the discovery of the Nodulation in absence of Rhizobium (also called Nar or spontaneous nodulation) in some genotypes of alfalfa (Medicago Sativa L.) (Truchet et al., 1989). This phenomenon demonstrates that Rhizobium not absolutely required for nodule organogenesis and raises many questions about the respective roles of the two partners. It also makes possible detailed analysis of the plant determinants involved in nodulation, avoiding any kind of exogenous activation.The ontogeny the spontaneous nodulation is similar to the Rhizobium-induced nodulation, beginning with cell divisions in the inner cortex (Joshi et al., 1991). The histological organisation of a fully developed spontaneous nodule is also characteristic of an indeterminate nodule, with an apical meristem, an outer cortex and an endodermis surrounding a central tissue and peripheral vascular bundles. Furthermore, the Nar phenomenon is i hibited by high concentrations of nitrogen in the culture medium and three early nodulins (i.e., nodule specific) genes (ENOD2, ENOD12 and have the same pattern of histological expression in spontaneous and ßhizobium-induced nodules. These data of ontology, histology, gene expression and nitrate regulation are consistent with the definition of a nodule, the main difference being the absence of any bacteria or infection thread and the presence of numerous starch grains within the central tissue. This last point led Caetano-Anollés et al. (1993) to hypothesize that nodules initially functioned as carbon-storage organs, later evolving to adapt to the Rhizobium nitrogen fixation. The spontaneous nodulation, observed in a small percentage (1-15%) all the cultivars of M. sativa tested so far, was recently described on Trifolíum repens, another allogamous, tetraploid legume species (Blauenfeldt et al., 1994). In alfalfa, the character is conserved during clonal propagation and is transmitted through meiosis (Truchet et al., 1989). A genetic model was proposed by Caetano-Anollès and Gresshoff (1 992). ' de des Plantes-Microorganismes, 27, 31 326 Castanet-Tolosan Tél. (33) 61 28 53 25 Fax: 61 28 50 61 [email protected] 177 CIHEAM Options Mediterraneennes