A New Bradyrhizobium japonicum Gene Required for Free-Living Growth and Bacteroid Development Is Conserved in Other Bacteria and in Plants. Marianne Weidenhaupt. Mikrobiologisches Institut, Eidgenossische Technische Hochschule, ETH-Zentrum, CH-8092 Zurich, Switzerland. Monika Schmid-Appert, BeatThony, Hauke Hennecke and Hans-Martin Fischer. Mikrobiologisches Institut, Eidgenossische Technische Hochschule, ETH-Zentrum, CH-8092 Zurich, Switzerland. MPMI 8: 454-464. Accepted 2 February 1995. Copyright 1995 The American Phytopathological Society.

In the nitrogen-fixing soybean symbiont Bradyrhizobium japonicum, a new DNA region, orf74, was discovered which is required for optimal free-living growth and, by consequence, also necessary for the formation of an effective symbiosis. A Tn5-233 insertion in orf74 resulted in a mutant, strain 74, that has a reduced growth rate in free-living cultures under all conditions tested and less than 1% residual symbiotic nitrogen fixation activity as compared with the wild type. Nodule distribution and nodule morphology are severely affected similarly as in a previously characterized B. japonicum nifA mutant. Protein databank searches revealed that the 142-amino-acid protein encoded by orf74 is homologous to a 161-amino-acid protein encoded by orf l 7 of Bacillus subtilis (approximately 46% identity; J. C. R. Struck, R. Kret-schmer-Kazemi Far, W. Schroder, F. Hucho, H. Y. Toschka, and V. A. Erdmann, Biochim. Biophys. Acta, 1050:80-83, 1990) as well as to a 178-amino-acid protein encoded by orfI78 of Escherichia coli (approximately 45% identity; K. L. Poulsen, N. W. Larsen, S. Molin, and P. Andersson, Mol. Microbiol., 6:895-905, 1992). Significant similarity was also found with unknown ORFs of two plant species. When expressed from a strong constitutive promoter, orfl 7 of B. subtilis could partially complement B. japonicum mutant 74. By contrast, orf74 of B. japonicum was unable to functionally complement an E. coli orfl78 mutant. The conservation of this DNA region in gram-negative and gram-positive bacteria suggests that the gene is essential for a fundamental cellular process which is required in B. japonicum for both free-living and symbiotic growth.