An ultrastructural and cytochemical study of the mature nodules from Trifolium repens L. indicates that bacteroid formation may be considered as a differentiation of the bacteria Rhizobium trifolii which, after invading the leguminous cell cytoplasm, undergo significant modifications including essentially: (a) progressive increase of the volume (about fortyfold), (b) modification of the shape of the bacteria which are successively short, rod-shaped, filamentous, pear-shaped, then spherical, and sometimes polyhedric, (c) dispersion of the initial nucleoid and appearance of granular bodies, (d) formation of a plasma membrane derived vesicular system, bearing or bounding oxydoreductive enzymes, which react with diaminobenzidine (DAB) at pH 6, 8, and 9. Bacteria to bacteroid transformation is related to host-cell differentiation which is mainly characterized by organelles distributed in concentric layers (peripheral starch-containing plastids, mitochondria, bacteroids, nucleus, and central vacuole) and by leghaemoglobin formation in the vacuolar space surrounding the bacteroids. Leghaemoglobin is detected by its electron density and positive reaction with DAB after incubation at alcaline pH in the presence of H2O2. The cytological events preceding or following the functional state of differentiation are also illustrated and discussed: infection of the host-cells by bacteria released from the threads, senescence of the bacteroids with formation of voluminous drops of poly-β-hydroxybutyrate (PHB), concomitant death of the host-cells, and the bacteroids at the base of the nodule.