Legume nodules from Lupinus albus L. cv. Multolupa and Phaseolus vulgaris L. cv. Tormesina plants were subjected to two different cryotechniques (High Pressure Freezing, HPF, and Fast Freeze Fixation, FFF) in order to compare the structure obtained with respect to previous features and to deepen the relationships between the microsymbiont and the host cell in the native state. Comparing both cryotechniques, HPF yielded better general nodule ultrastructure than FFF. HPF allowed the observation of the native nodule structure since it permitted an excellent preservation. The result obtained showed that the peribacteroidal membrane was in cióse contact with the bacteroids, which is in contrast with the observations after chemical fixation. The narrow peribacteroid space observed between bacteroid and peribacteroid membranes appears as a dense matrix which is in agreement with the former knowledge on the existence of some enzymes and proteins in this space. CVreleased by leghemoglobin will occur in cióse proximity to the bacteroid membranes. The spatial relationship between Golgi apparatus and ER with the peribacteroidal membrane observed along the development of the nodule indicates that these structures are very much related with the formation of the symbiosome membrane. HPF could be the right technique to continué studying real functional aspects in developing nodules.

The HPF method was performed with a HPM 010 Bal-Tec (Licchtenstein) machine at the Centre Interuniversitaire de Microscopie Electronique (CIME-Jussieu) in Paris (France). We thank Ghislaine Frébourg for her expert technical assistance for the preparation fo the sandwiched specimens for HPF. We also thank Carmen de Mesa and Maria Isabel Menendez for competent and skillful technical assistance and to Fernando Pinto from the Electron Microscope Service for facilities.

9 pages, figures, and tables statistics.

Peer reviewed