Application of 1-aminoocyclopropane-1-carboxylic acid, an ethylene precursor, decreased nodulation of Macroptilium atropurpureum by Bradyrhizobium elkanii. B. elkanii produces rhizobitoxine, an ethylene synthesis inhibitor. Elimination of rhizobitoxine production in B. elkanii increased ethylene evolution and decreased nodulation and competitiveness on M. atropurpureum. These results suggest that rhizobitoxine enhances nodulation and competitiveness of B. elkanii on M. atropurpureum.The symbiotic interactions between a legume and (brady) rhizobia result in a unique, nitrogen-fixing plant organ, the nodule. Recent studies have shown that the phytohormone ethylene inhibits nodule formation in some legumes (8,9,16,24,25). Application of 1-aminoocyclopropane-1-carboxylic acid (ACC), a precursor of ethylene, inhibits nodulation in Medicago truncatula (24).Rhizobitoxine [2-amino-4-(2-amino-3-hydropropoxy)-transbut-3-enoic acid] is an ethylene synthesis inhibitor that is produced by the legume symbiont Bradyrhizobium elkanii (15, 17-19, 22, 39). It is thought that production of this compound enhances nodulation of the host legume because of its inhibitory effect on ethylene synthesis. However, some reports have shown that there is not a significant difference in nodule number between plants inoculated with B. elkanii USDA61 and plants inoculated with rhizobitoxine-deficient mutants during nodulation of Glycine max, Glycine soja, Vigna unguiculata, and Macroptilium atropurpureum (26,39). Recently, Duodu et al. observed a significant difference in nodule number between plants inoculated with isogenic variants of USDA61 during nodulation of Vigna radiata (7). Although these findings do not seem to be consistent with the hypothesis that rhizobitoxine has a positive effect on nodulation, the inconsistency can be explained by differences in the ethylene sensitivity of nodulation among leguminous species; nodulation of G. max is generally not sensitive to ethylene (10, 31, 38), while nodulation of V. radiata is sensitive (7). The inconsistency could also result from differences in the abilities of the strains used in the experiments to produce rhizobitoxine; strain USDA61 is a weak producer of rhizobitoxine (39).In addition to G. max, the leguminous plant M. atropurpureum is a nodulating host for B. elkanii and Bradyrhizobium japonicum (12,15). Although the effect of ethylene on nodulation has been studied in many leguminous host plants so far, the effect of ethylene in M. atropurpureum is not known. B. elkanii was found to be more competitive than B. japonicum for nodulation of M. atropurpureum in a multistrain environment when a field soil was inoculated with a mixture of several strains isolated from the field soil (21). In general, B. elkanii accumulates rhizobitoxine in cultures and in nodules, while B. japonicum does not (5,15,18,19). These results led us to investigate the role of rhizobitoxine production on the nodulation and competitiveness of B. elkanii on M. atropurpureum by using a B. elkanii strain that produces...

In a survey of DNA fingerprints of indigenous Bradyrhizobium japonicum with the species-specific repeated sequences RSα and RSβ, 21 isolates from three field sites showed numerous RS-specific hybridization bands. The isolates were designated highly reiterated sequence-possessing (HRS) isolates, and their DNA hybridization profiles were easily distinguished from the normal patterns. Some HRS isolates from two field sites possessed extremely high numbers of RSα copies, ranging from 86 to 175 (average, 128), and showed shifts and duplications of nif- and hup-specific hybridization bands. The HRS isolates exhibited slower growth than normal isolates, although no difference in symbiotic properties was detected between the HRS and normal isolates. Nucleotide sequence analysis of 16S rRNA genes showed that HRS isolates were strains ofB. japonicum. There was no difference in the spectra of serological and hydrogenase groupings of normal and HRS isolates. Some HRS isolates possessed a tandem repeat RSα dimer that is similar to the structure of (IS30)2, which was shown to cause a burst of transpositional rearrangements in Escherichia coli. The results suggest that HRS isolates are derived from normal isolates in individual fields by genome rearrangements that may be mediated by insertion sequences such as RSα.

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