Figure 4

Rhizobial differentiation in Fix- mutants. Rhizobial colonization and bacterial morphology in M. truncatula wild-type (A-C) and Fix- mutant nodules (D-O). Longitudinal sections of nodules 4 wpi with S. meliloti were stained with SYTO13 and analyzed by confocal microscopy. The regions of the mature wild-type (wt) nodules (A) indicated similarly as in Figure 2; but also with IV., senescence zone. Higher magnification revealed elongated bacteroids in the infected cells of the interzone and the nitrogen-fixation zone of wild type nodules (B and C). The reduced zonation phenotype of dnf5-2 (D) is coupled with the lack of bacterial elongation (G). The bacterial development is initiated in the nodules of the other ineffective mutants (5L, E and H; 11S, F and I; 7Y, J and M; 13U, K and N; dnf8, L and O). Strong autofluorescence was observed in the proximal part of zone III of 7Y nodules (J). Violet-laser excited (405 nm) autofluorescence was detected between 545–645 nm and pseudocolored in red (A and J). Arrow shows an infection thread in panel M, arrowhead indicates branched bacteroids in panel N. Bars represent 200 μm in A, D, E, F, J, K and L, 20 μm in panels B, C, G, H, I, M, N and O.