Aims:  In order to depict the fine interactions that lead to nodulation, absolute microbiological control of the symbiotic partners is required, i.e. the ability to obtain in vitro axenic nodulation, a condition that has never been fulfilled with the Casuarina–Frankia symbiosis. The effects of culture conditions on plant growth and nodule formation by Casuarina cunninghamiana were investigated. Methods and Results:  Axenic (capped tubes with different substrates), and nonaxenic cultures (Gibson tubes, pot cultures) were tested. In axenic conditions, C. cunninghamiana, inoculated with Frankia, had poor growth and did not form nodules at 6 weeks. Plants cultivated in Gibson tubes reached the four axillary shoots stage within 6 weeks and formed nodules 4 weeks after inoculation. Sand‐pot cultures allowed us to relate the plant development stage at inoculation with nodulation. Conclusions:  The sterile replacement of the cap by a plastic bag increased plant growth and enabled nodule formation 6 weeks after inoculation. The new system of plant culture allows the axenic nodule formation 6 weeks after inoculation. Nodulation behaviour is related to plant development and confinement. Significance and Impact of the Study:  This axenic plant nodulation system is of major interest in analysing the roles of Frankia genes in nodulation pathways.

Abstract.-The formation of Casuarina actinorhizae in axenic conditions remains a challenge hindering research program. To explore the putative role of helper bacteria favouring actinorhizae formation, 50 bacteria were tested in an original selection scheme for their effects on C. cunninghamiana growth in the presence of Frankia. Two strains FAB1 and FAB2 were identified as having a significant promoting effect on C. cunninghamiana growth in the presence of Frankia. Actinorhizae formation was never observed in these strictly axenic conditions. The comparison of 165 rDNA sequences shows similarity of 99% for FAB1 with Bacillus sp. and of 99% for FAB2 with Ochrobactrum sp.Key words : axenic culture -Casuarina -Frankia -helper bacteria -symbiosis.