Diversity of new root nodule bacteria from Erythrina velutina Willd., a native legume from the Caatinga dry forest (Northeastern Brazil).

This study aims to evaluate the phenotypical characteristics of bacterial isolates from mulungu (Erythrina velutina Willd.) nodules and determinate their Box-PCR fingerprinting. All bacteria were evaluated by the following phenotypic characteristics: growth rate, pH change, colony color and mucus production. The bacterial isolates able to re-nodulate the original host were also evaluated regarding its tolerance to increased salinity and different incubation temperatures, ability to growth using different carbon sources, intrinsic antibiotic resistance and ?in vitro? auxin biosynthesis. The molecular fingerprints were set up using the Box-PCR technique and the isolates were clustered by their profiles. Among the 22 bacterial isolates obtained, eight were able to re-nodulate the original host. Among the nodule inducing isolates, some were tolerant to 1% of NaCl and 39° C and all of them metabolized the maltose, fructose, glucose, sucrose and arabinose, were resistant to rifampicin and produced auxin. The bacteria showed low genetic similarity among them and reference strains, which indicates the great genetic variability of the isolates. The results of this work are the first reports about the bacterial isolates able to nodulate this species. A more deep study of these bacteria may reveal the existence of isolates tolerant to environmental stresses and suitable as a future mulungu inoculant.

Comparative genomics suggests that an ancestral polyploidy event leads to enhanced root nodule symbiosis in the Papilionoideae

Root nodule symbiosis (RNS) is one of the most efficient biological systems for nitrogen fixation and it occurs in 90% of genera in the Papilionoideae, the largest subfamily of legumes. Most papilionoid species show evidence of a polyploidy event occurred approximately 58 million years ago. Although polyploidy is considered to be an important evolutionary force in plants, the role of this papilionoid polyploidy event, especially its association with RNS, is not understood. In this study, we explored this role using an integrated comparative genomic approach and conducted gene expression comparisons and gene ontology enrichment analyses. The results show the following: (1) approximately a quarter of the papilionoid-polyploidy-derived duplicate genes are retained; (2) there is a striking divergence in the level of expression of gene duplicate pairs derived from the polyploidy event; and (3) the retained duplicates are frequently involved in the processes crucial for RNS establishment, such as symbiotic signalling, nodule organogenesis, rhizobial infection and nutrient exchange and transport. Thus, we conclude that the papilionoid polyploidy event might have further refined RNS and induced a more robust and enhanced symbiotic system. This conclusion partly explains the widespread occurrence of the Papilionoideae.

REMEDIATION OF TETRACYCLINE FROM WATER SOURCES USING VETIVER GRASS (Chrysopogon zizanioides L. Nash) AND TETRACYCLINE-TOLERANT ROOT-ASSOCIATED BACTERIA

Antibiotics are emerging contaminants worldwide. Due to insufficient policy regulations, public awareness, and the constant exposure of the environment to antibiotic sources has created a major environmental concern. Wastewater treatment plants (WWTP) are not equipped to filter-out these compounds before the discharge of the disinfected effluent into water sources (e.g., lakes and streams) and current available technologies are not equipped to remediate these compounds from environmental sources. Hence, the challenge remains to establish a biological system to remove these antibiotics from wastewater. An invitro hydroponic remediation system was developed using vetiver grass (Chrysopogon zizanioides L. Nash) to remediate tetracycline (TC) from water. Comparative metabolomics studies were conducted to investigate the metabolites/pathways associated with tetracycline metabolism in plants and TC-degrading bacteria. The results show that vetiver plants effectively uptake tetracycline from water sources. Vetiver root-associated bacteria recovered during the hydroponic remediation trial were highly tolerant to TC (as high as 600 ppm) and could use TC as a sole carbon and energy source. Growth conditions (pH, temperature, and oxygen requirement) for TC-tolerant bacteria were optimized for higher TC remediation capability from water sources. The plant (roots and shoots) and bacterial species were further characterized for the metabolites produced during the TC degradation process using GC-MS to identify the possible biochemical mechanism involved. Also, the plant root zone was screened for metabolites/enzymes that were secreted during antibiotic degradation and could potentially enhance the degradation process. The root zone was selected for this analysis because this region of the plant has shown a greater capacity for antibiotic degradation compared to the shoot zone. The role of antioxidant enzymes in TC degradation process revealed glutathione-S-transferase (GSTs) as an important group of enzymes in both plant and bacteria potentially involved in TC degradation process. Metabolomics results also suggest potential GST activity in the TC remediation/ transformation process used by plants. This information could be useful in gaining insights for the application of biological remediation systems for the mitigation of antibiotics from waste-water.

Ethylene-mediated phenotypic plasticity in root nodule development on Sesbania rostrata

Leguminous plants in symbiosis with rhizobia form either indeterminate nodules with a persistent meristem or determinate nodules with a transient meristematic region. Sesbania rostrata was thought to possess determinate stem and root nodules. However, the nature of nodule development is hybrid, and the early stages resemble those of indeterminate nodules. Here we show that, depending on the environmental conditions, mature root nodules can be of the indeterminate type. In situ hybridizations with molecular markers for plant cell division, as well as the patterns of bacterial nod and nif gene expression, confirmed the indeterminate nature of 30-day-old functional root nodules. Experimental data provide evidence that the switch in nodule type is mediated by the plant hormone ethylene.

Oxygen as a key developmental regulator of Rhizobium meliloti N2-fixation gene expression within the alfalfa root nodule.

The symbiotic pattern of expression of Rhizobium meliloti N2-fixation genes is tightly coupled with the histological organization of the alfalfa root nodule and thus is under developmental control. N2-fixation gene expression is induced very sharply at a particular zone of the nodule called interzone II-III that precedes the zone where N2 fixation takes place. We show here that this coupling can be disrupted, hereby resulting in ectopic expression of N2-fixation genes in the prefixing zone II of the nodule. Uncoupling was obtained either by using a R. meliloti strain in which a mutation rendered N2-fixation gene expression constitutive with respect to oxygen in free-living bacterial cultures or by placing nodules induced by a wild-type R. meliloti strain in a microoxic environment. These results implicate oxygen as a key determinant of the symbiotic pattern of N2-fixation gene expression.

Complete genome sequence of the Medicago microsymbiont Ensifer (Sinorhizobium) medicae strain WSM419

Ensifer (Sinorhizobium) medicae is an effective nitrogen fixing microsymbiont of a diverse range of annual Medicago (medic) species. Strain WSM419 is an aerobic, motile, non-spore forming, Gram-negative rod isolated from a M. murex root nodule collected in Sardinia, Italy in 1981. WSM419 was manufactured commercially in Australia as an inoculant for annual medics during 1985 to 1993 due to its nitrogen fixation, saprophytic competence and acid tolerance properties. Here we describe the basic features of this organism, together with the complete genome sequence, and annotation. This is the first report of a complete genome sequence for a microsymbiont of the group of annual medic species adapted to acid soils. We reveal that its genome size is 6,817,576 bp encoding 6,518 protein-coding genes and 81 RNA only encoding genes. The genome contains a chromosome of size 3,781,904 bp and 3 plasmids of size 1,570,951, 1,245,408 and 219,313 bp. The smallest plasmid is a feature unique to this medic microsymbiont.

Complete genome sequence of Rhizobium leguminosarum bv trifolii strain WSM2304, an effective microsymbiont of the South American clover Trifolium polymorphum

Rhizobium leguminosarum bv. trifolii is the effective nitrogen fixing microsymbiont of a diverse range of annual and perennial Trifolium (clover) species. Strain WSM2304 is an aerobic, motile, non-spore forming, Gram-negative rod isolated from Trifolium polymorphum in Uruguay in 1998. This microsymbiont predominated in the perennial grasslands of Glencoe Research Station, in Uruguay, to competitively nodulate its host, and fix atmospheric nitrogen. Here we describe the basic features of WSM2304, together with the complete genome sequence, and annotation. This is the first completed genome sequence for a nitrogen fixing microsymbiont of a clover species from the American centre of origin. We reveal that its genome size is 6,872,702 bp encoding 6,643 protein-coding genes and 62 RNA only encoding genes. This multipartite genome was found to contain 5 distinct replicons; a chromosome of size 4,537,948 bp and four circular plasmids of size 4,537,948, 1,266,105, 501,946, 308,747 and 257,956 bp.

Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain SRDI565

Rhizobium leguminosarum bv. trifolii SRDI565 (syn. N8-J) is an aerobic,motile, Gram-negative, non-spore-forming rod. SRDI565 was isolated from anodule recovered from the roots of the annual clover Trifolium subterraneum subsp. subterraneum grown in thegreenhouse and inoculated with soil collected from New South Wales, Australia. SRDI565has a broad host range for nodulation within the clover genus, however N2-fixationis sub-optimal with some Trifoliumspecies and ineffective with others. Here we describe the features of R. leguminosarum bv. trifolii strain SRDI565, together with genomesequence information and annotation. The 6,905,599 bp high-quality-draft genomeis arranged into 7 scaffolds of 7 contigs, contains 6,750 protein-coding genesand 86 RNA-only encoding genes, and is one of 100 rhizobial genomes sequencedas part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia forBacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain SRDI943

Rhizobium leguminosarum bv. trifolii SRDI943(syn. V2-2) is an aerobic, motile, Gram-negative, non-spore-forming rod. SRDI943was isolated from a nodule recovered from the roots of the annual clover Trifoliummichelianum savi cv. paradanathat had been inoculated with a soil collected from a mixed pasture in Victoria, Australia. SRDI943 has a broadhost range for nodulation within the clover genus, however N2-fixationis sub-optimal (20-54% of reference strain WSM1325) on T. subterraneum spp.Here we describe the features of R. leguminosarum bv. trifolii strain SRDI943, together with genomesequence information and annotation. The 7,412,387 bp high-quality-draft genomeis arranged into 5 scaffolds of 5 contigs, contains 7,317 protein-coding genesand 89 RNA-only encoding genes, and is one of 100 rhizobial genomes sequencedas part of the DOE Joint Genome Institute 2010 Genomic Encylopedia for Bacteriaand Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Genome sequence of Ensifer sp. TW10, a Tephrosia wallichii (Biyani) microsymbiont native to the Indian Thar Desert

Ensifer sp. TW10 is novel N2-fixingbacterium isolated from a root nodule of the perennial legume Tephrosia wallichii Graham (known locally as Biyani) found in the Great Indian (or Thar) desert, a large arid regionin the northwestern part of the Indian subcontinent. Strain TW10 is a Gram-negative, rod shaped,aerobic, motile, non-spore forming, species of root nodule bacteria (RNB) that promiscuously nodulates legumes in Thar Desert alkaline soil. It is fast growing, acid-producing, and tolerates up to 2% NaCl and capable of growth at 40C. In this report we describe for the first time the primary features of this Thar Desert soil saprophyte together with genome sequence informationand annotation. The 6,802,256 bp genome has a GC content of 62% and is arranged into 57 scaffolds containing 6,470 protein-coding genes, 73 RNA genes and asingle rRNA operon. This genome is one of 100 RNB genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Genome sequence of Ensifer medicae strain WSM1369, an effective microsymbiont of the annual legume Medicago sphaerocarpos

Ensifer medicae WSM1369 is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of Medicago. WSM1369 was isolated in 1993 from a nodule recovered from the roots of Medicago sphaerocarpos growing at San Pietro di Rudas, near Aggius in Sardinia (Italy). WSM1369 is an effective microsymbiont of the annual forage legumes M. polymorpha and M. sphaerocarpos. Here we describe the features of E. medicae WSM1369, together with genome sequence information and its annotation. The 6,402,557 bp standard draft genome is arranged into 307 scaffolds of 307 contigs containing 6,656 protein-coding genes and 79 RNA-only encoding genes. This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Genome sequence of Ensifer medicae strain WSM1115, an acid-tolerant Medicago-nodulating microsymbiont from Samothraki, Greece

Ensifer medicae (syn. Sinorhizobium medicae) strain WSM1115 forms effective nitrogen fixing symbioses with a range of annual Medicago species and is used in commercial inoculants in Australia. WSM1115 is an aerobic, motile, Gram-negative, non-spore-forming rod. It was isolated from a nodule recovered from the root of burr medic (Medicago polymorpha) collected on the Greek Island of Samothraki. WSM1115 has a broad host range for nodulation and N2 fixation capacity within the genus Medicago, although this does not extend to all medic species. WSM1115 is considered saprophytically competent in moderately acid soils (pHCaCl2 5.0) however, has failed to persist at field sites where soil salinity exceeded 10 ECe (dS/m). Here we describe the features of E. medicae strain WSM1115, together with genome sequence information and its annotation. The 6,861,065 bp high-quality-draft genome is arranged into 7 scaffolds of 28 contigs, contains 6,789 protein-coding genes and 83 RNA-only encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Genome sequence of Ensifer arboris strain LMG 14919T, a microsymbiont of the legume Prosopis chilensis growing in Kosti, Sudan

Ensifer arboris LMG 14919T is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of several species of legume trees. LMG 14919T was isolated in 1987 from a nodule recovered from the roots of the tree Prosopis chilensis growing in Kosti, Sudan. LMG 14919T is highly effective at fixing nitrogen with P. chilensis (Chilean mesquite) and Acacia senegal (gum Arabic tree or gum acacia). LMG 14919T does not nodulate the tree Leucena leucocephala, nor the herbaceous species Macroptilium atropurpureum, Trifolium pratense, Medicago sativa, Lotus corniculatus and Galega orientalis. Here we describe the features of E. arboris LMG 14919T, together with genome sequence information and its annotation. The 6,850,303 bp high-quality-draft genome is arranged into 7 scaffolds of 12 contigs containing 6,461 protein-coding genes and 84 RNA-only encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.