Improved soybean root association of N-starved Bradyrhizobium japonicum

LÓPEZ-GARCÍA, S.L.; VÁZQUEZ, T.E.E.; FAVELUKES, G.; LODEIRO, A.R.

JOURNAL OF BACTERIOLOGY

American Society of Microbiology

Año: 2001 vol. 183 p. 7241 - 7252

0021-9193

Despite the well known effects of combined nitrogen on the N-fixing symbiosis between Rhizobiaceae and legumes, relatively few studies have addressed the consequences of N-limitation in the rhizobia in its association with the plant root. In this study, we observed that Bradyrhizobium japonicum is able to scavenge very low amounts of N from its surroundings: the wild type strain LP 3001 grew for six generations with a growth rate of 1.2 day-1 in a Götz minimal medium (Götz, R. et al., 1982. J. Gen. Microbiol. 128:789-798) with 28 mM mannitol as carbon source, and with the N-source [(NH4)2SO4] limited to only 20 M. Under these conditions the glutamine synthetase (GS) activity, which has a key role in N assimilation, was 5-6 times higher than in similar cultures grown with 1 mM (NH4)2SO4 or 0.1 mM (NH4)2SO4. The NtrBC-inducible GS II form of this enzyme accounted for 60% of the specific activity in N-starved rhizobia, being negligible in the other two cultures. The exopolysaccharide (EPS) and capsular polysaccaride (CPS) contents relative to cell protein were significantly higher in the N-starved cultures, but on the other hand, the poly-3-hydroxybutyrate did not rise in comparison with N-sufficient cultures. Early interactions of rhizobia with soybean lectin (SBL) or with flavonoids, as well as nodulation efficiency and competitiveness for nodule occupation, were also affected by N-starvation. In agreement with the accumulation of CPS in N-starved cultures, SBL binding as well as stimulation of rhizobial adsorption to soybean roots by SBL pretreatment, were higher. The last effect was evident only in cultures that had not entered stationary phase. In stationary phase cultures, adsorption was not stimulated by lectin treatment independently of their N-limitation state, CPS content, or SBL binding activity. We also studied nodC gene induction in relation to N-starvation. In the chromosomal nodC::lacZ fusion Bj110-573, nodC gene expression was induced by genistein 2.7-fold more in N-starved young cultures than in non-starved ones. In stationary cultures, nodC gene expression was similarly induced in N-limited cultures but induction was negligible in cultures limited by another nutrient. Nodulation profiles obtained with strain LP 3001 grown under N-starvation indicated that these cultures nodulated faster. In addition, as culture age increased, nodulation efficiency decreased for two reasons: less nodules were formed and nodulation was delayed. However, their relative importance was different according to the nutrient condition: the overall decrease in the number of nodules in older cultures was the main effect in N-starved cultures whereas a delay in nodulation was more responsible for a loss in efficiency of N-sufficient broths. Competition for nodulation was studied with young cultures of two wild type strains differing only in their antibiotic resistance, the N-starved cultures being the most competitive for nodule occupation.