Diversity of Sinorhizobium (Ensifer) meliloti Bacteriophages in the Rhizosphere of Medicago marina: Myoviruses, Filamentous and N4-Like Podovirus.

Using different Sinorhizobium meliloti strains as hosts, we isolated eight new virulent phages from the rhizosphere of the coastal legume Medicago marina. Half of the isolated phages showed a very narrow host range while the other half exhibited a wider host range within the strains tested. Electron microscopy studies showed that phages M_ort18, M_sf1.2, and M_sf3.33 belonged to the Myoviridae family with feature long, contractile tails and icosaedral head. Phages I_sf3.21 and I_sf3.10T appeared to have filamentous shape and produced turbid plaques, which is a characteristic of phages from the Inoviridae family. Phage P_ort11 is a member of the Podoviridae, with an icosahedral head and a short tail and was selected for further characterization and genome sequencing. P_ort11 contained linear, double-stranded DNA with a length of 75239 bp and 103 putative open reading frames. BLASTP analysis revealed strong similarities to Escherichia phage N4 and other N4-like phages. This is the first report of filamentous and N4-like phages that infect S. meliloti.

RNA-seq analysis of the Rhizobium tropici CIAT 899 transcriptome shows similarities in the activation patterns of symbiotic genes in the presence of apigenin and salt.

BACKGROUND: Rhizobium tropici strain CIAT 899 establishes effective symbioses with several legume species, including Phaseolus vulgaris and Leucaena leucocephala. This bacterium synthesizes a large variety of nodulation factors in response to nod-gene inducing flavonoids and, surprisingly, also under salt stress conditions. The aim of this study was to identify differentially expressed genes in the presence of both inducer molecules, and analyze the promoter regions located upstream of these genes. RESULTS: Results obtained by RNA-seq analyses of CIAT 899 induced with apigenin, a nod gene-inducing flavonoid for this strain, or salt allowed the identification of 19 and 790 differentially expressed genes, respectively. Fifteen of these genes were up-regulated in both conditions and were involved in the synthesis of both Nod factors and indole-3-acetic acid. Transcription of these genes was presumably activated through binding of at least one of the five NodD proteins present in this strain to specific nod box promoter sequences when the bacterium was induced by both apigenin and salt. Finally, under saline conditions, many other transcriptional responses were detected, including an increase in the transcription of genes involved in trehalose catabolism, chemotaxis and protein secretion, as well as ribosomal genes, and a decrease in the transcription of genes involved in transmembrane transport. CONCLUSIONS: To our knowledge this is the first time that a transcriptomic study shows that salt stress induces the expression of nodulation genes in the absence of flavonoids. Thus, in the presence of both nodulation inducer molecules, apigenin and salt, R. tropici CIAT 899 up-regulated the same set of symbiotic genes. It could be possible that the increases in the transcription levels of several genes related to nodulation under saline conditions could represent a strategy to establish symbiosis under abiotic stressing conditions.

Mutation in the cobO gene generates auxotrophy for cobalamin and methionine and impairs the symbiotic properties of Sinorhizobium fredii HH103 with soybean and other legumes.

We report here the isolation of a methionine and cobalamin mutant strain (SVQ336) of Sinorhizobium fredii HH103 obtained by Tn5-lacZ mutagenesis. Sequence analysis showed that the transposon was inserted into a gene homologous to cobO. This gene codes for a cobalamin adenosyltransferase which is involved in the biosynthesis of vitamin B12. Another HH103 cobO mutant (strain SVQ524), was constructed by the insertion of Omega interposon. Both cobO mutants required the addition of methionine because cobalamin acts as a cofactor of the enzyme MetH, which catalyses the last step of the methionine biosynthesis. Mutant SVQ524 failed to nodulate on Vigna radiate but was able to nodulate on Glycine max cvs. Williams and Peking and Cajanus cajan, although the total number of nodules formed was highly reduced in comparison with that of plants inoculated with the wild-type strain HH103. The roots of these plants did not seem to secrete enough cobalamin and/or methionine to support growth of cobalamin/methionine auxotrophs in the rhizosphere. In all cases, the phenotype of SVQ524 was nearly overcome by the addition of methionine or cobalamin to the plant growth media or by the presence of a copy of the cobO gene in cosmid pMUS756.

Inactivation of the Sinorhizobium fredii HH103 rhcJ gene abolishes nodulation outer proteins (Nops) secretion and decreases the symbiotic capacity with soybean.

It has been postulated that nodulation outer proteins (Nops) avoid effective nodulation of Sinorhizobium fredii USDA257 to nodulate with American soybeans. S. fredii HH103 naturally nodulates with both Asiatic (non-commercial) and American (commercial) soybeans. Inactivation of the S. fredii HH103 gene rhcJ, which belongs to the tts (type III secretion) cluster, abolished Nop secretion and decreased its symbiotic capacity with the two varieties of soybeans. S. fredii strains HH103 and USDA257, that only nodulates with Asian soybeans, showed different SDS-PAGE Nop profiles, indicating that these strains secrete different sets of Nops. In coinoculation experiments, the presence of strain USDA257 provoked a clear reduction of the nodulation ability of strain HH103 with the American soybean cultivar Williams. These results suggest that S. fredii Nops can act as either detrimental or beneficial symbiotic factors in a strain-cultivar-dependent manner. Differences in the flavonoid-mediated expression of rhcJ with respect to nodA were also detected. In addition, one of the Nops secreted by strain HH103 was identified as NopA.

Inactivation of the Sinorhizobium fredii HH103 rhcJ gene abolishes nodulation outer proteins (Nops) secretion and decreases the symbiotic capacity with soybean / La inactivación del gen rhcJ de Sinorhizobium fredii HH103 elimina la secreción de las proteínas externas de nodulación (Nops) y disminuye la capacidad de simbiosis con la soja

It has been postulated that nodulation outer proteins (Nops) avoid effective nodulation of Sinorhizobium fredii USDA257 to nodulate with American soybeans. S. fredii HH103 naturally nodulates with both Asiatic (non-commercial) and American (commercial) soybeans. Inactivation of the S. fredii HH103 gene rhcJ, which belongs to the tts (type III secretion) cluster, abolished Nop secretion and decreased its symbiotic capacity with the two varieties of soybeans. S. fredii strains HH103 and USDA257, that only nodulates with Asian soybeans, showed different SDS-PAGE Nop profiles, indicating that these strains secrete different sets of Nops. In coinoculation experiments, the presence of strain USDA257 provoked a clear reduction of the nodulation ability of strain HH103 with the American soybean cultivar Williams. These results suggest that S. fredii Nops can act as either detrimental or beneficial symbiotic factors in a strain-cultivar-dependent manner. Differences in the flavonoid-mediated expression of rhcJ with respect to nodA were also detected. In addition, one of the Nops secreted by strain HH103 was identified as NopA (AU)

Se ha propuesto que las proteínas externas de nodulación (Nops) impiden la nodulación efectiva de Sinorhizobium fredii USDA257 con las sojas americanas. S. fredii HH103 nodula de forma natural tanto con las sojas asiáticas (no comercializadas) como con las americanas (comercializadas). La inactivación del gen rhcJ de HH103, que pertenece a la agrupación génica tts (secreción de tipo III), anuló la secreción de Nops y redujo la capacidad simbiótica de esta bacteria con las dos variedades de soja. Las cepas HH103 y USDA257 de S. fredii, que sólo nodula sojas asiáticas, mostraron perfiles SDS-PAGE diferentes de Nop, lo cual sugiere que estas cepas podrían secretar distintos conjuntos de Nops. Cuando las cepas USDA257 y HH103 fueron inoculadas conjuntamente, la capacidad de nodulación de esta última cepa con el cultivar americano Williams de soja se redujo significativamente. Estos resultados indican que las Nops secretadas por S. fredii pueden actuar como factores simbióticos tanto positivos como negativos dependiendo de la cepa-cultivar rizobiana. Se detectaron también diferencias entre la expresión mediada por flavonoides del gen rhcJ y del nodA. Además, una de las Nops secretadas por la cepa HH103 fue identificada como NopA (AU)