Nitrogen nutrition and the development and senescence of nodules on cowpea seedlings

Cowpea (Vigna unguiculata (L.) Walp cv. Vita 3) seedlings inoculated with Rhizobium strain CB756 were cultured with their root systems maintained in air or in Ar:O2 (80:20, v/v) during early nodule development (up to 24 d after sowing). Compared with those in air, seedlings in Ar:O2 showed progressive N deficiency with inhibited shoot growth, reduced ribulose-1,5-bisphosphate carboxylase and total protein levels and loss of chlorophyll in the leaves. Nodule initiation, differentiation of infected and uninfected nodule tissues and the ultrastructure of bacteroid-containing cells were similar in the air and Ar:O2 treatments up to 16 d after sowing. Thereafter the Ar:O2 treatment caused cessation of growth and development of nodules, reduced protein levels in bacteroids and nodule plant cells, and progressive degeneration of nodule ultrastructure leading to premature senescence of these organs. Provision of $\mathrm{N}{\mathrm{O}}_{3}^{-}$ (0.1—0.2 mM) to Ar:O2-grown seedlings overcame the abovementioned consequences of N2 deficiency on nodule and plant growth, but merely delayed the degenerative effects of Ar:O2 treatment on nodule structure and senescence. Treatment of Ar:O2-grown seedlings with $\mathrm{N}{\mathrm{O}}_{3}^{-}$ greatly increased the protein level of nodules but the increase was largely restricted to the plant cell fraction as opposed to the bacteroids. By contrast, $\mathrm{N}{\mathrm{O}}_{3}^{-}$ treatment of air-grown seedlings increased protein of bacteroid and host nodule fractions to the same relative extents when compared with air-grown plants not supplemented with $\mathrm{N}{\mathrm{O}}_{3}^{-}$. These findings, taken together with studies of the distribution of N in nodules of symbiotically effective plants grown from 15N-labeled seed, indicate that direct incorporation of fixation products by bacteroids may be a critical feature in the establishment and continued growth of an effective symbiosis in the cowpea seedling.