Partial purification of a competence factor from Rhizobium japonicum

A competence factor (CF) from Rhizobium japonicum was partially purified to 43 fold on Sephadex G-100. This CF preparation was sensitive to heat, trypsin and pronase, was resistant to DNase 1, RNase A and lysozyme. It had an approximate mol. wt. of 82,000. Osmotic shock treatment of competent cells revealed that the CF is located in the periplasmic region of the cell.Abbreviations CF competence factor - BSA bovine serum albumin - YM yeast mannitol medium     

Studies on transducing phage M-1 for Rhizobium japonicum D211

Phage M-1 produced clear plaques with a halo in the lawn of Rhizobium japonicum D211. A one step growth curve of phage M-1 showed a latent period of 3 h, burst size of 55 and rise period of 2 h. The inactivation of phage M-1 was found to be dependent upon the concentraion of d-glucosomanine. The neutralization kinetics of phage M-1 by antiphage serum gave a K value (velocity constant) of 83.1 min^–1. Transduction of str and kan was studied in the presence of antiphage serum and d-glucosamine. Cotransduction of different antibiotic resistance markers suggested that the system can be further explored for high resolution mapping in R. japonicum.Abbreviations YM yeast mannitol medium - PFU plaque forming unit - moi multiplicity of infection - EOP efficiency of plating     

Plants interact with microbial polysaccharides

Plants are resistant to almost all of the microorganisms with which they come in contact. In response to invasion by a fungus, bacterium, or a virus, many plants produce low molecular weight compounds, phytoalexins, which inhibit the growth of microorganisms. Phytoalexins are produced whether or not the invading microorganism is a pathogen. The production of phytoalexins appears to be a widespread mechanism by which plants attempt to defend themselves against pests. Molecules of microbial origin which trigger phytoalexin accumulation in plants are called elicitors. Structural polysaccharides from the mycelial walls of several fungi elicit phytoalexin accumlation in plants. Approximately 10 ng of the polysaccharide elicits the accumulation in plants of more than sufficient amounts of phytoalexin to stop the growth of microorganisms in vitro. The best characterized elicitors have been demonstrated to be β-1,3-glucans with branches to the 6 position of some of the glucosyl residues. Oligosaccharides, produced by partial acid hydrolysis of the mycelial wall glucans, are exceptionally active elicitors. The smallest oligosaccharide which is still an effective elicitor is composed of about 8 sugar residues. Bacteria also elicit phytoalexin accumulation in plants, but the Rhizobium symbionts of legumes presumably have a mechanism which allows them to avoid either eliciting phytoalexin accumulation or the effects of the phytoalexins if they are accumulated. The lectins of legumes bind to the lipopolysaccharides of their symbiont, but not of their non-symbiont, Rhizobium. It is not known whether the lectin-lipopolysaccharide interaction is involved with the establishment of symbiosis. However, evidence will be presented that suggests that lectins are, in fact, enzymes capable of modifying the structurs of the lipopolysaccharides of their symbiont, but not of their non-symbiont, Rhizobium. It will also be shown that the lipopolysaccharides isolated from different Rhizobium species and from different strains of individual Rhizobium species have different sugar compositions. Thus, the different strains of a single Rhizobium species are as different from one another as the different species of Salmonella and other gram-negative bacteria. This conclusion is substantiated by experiments demonstrating that antibodies to the lipopolysaccharide from a single Rhizobium strain can differentiate that strain from other strains of the same species as well as from other Rhizobium species. The role in symbiosis of the strain-specific O-antigens is unknown.     

The relation between nitrogen percent and dry weight of inoculated legumes

Summary Results of numerous tests of tropical and temperate legume hosts and Rhizobium strains accumulated between 1952 and 1966 were examined for a relation between N percent (y) and dry weight per plant (x). The data fitted the equation y=A–be^–cx. The most effective Rhizobium strains can be selected on the basis of dry matter yields of whole plants or plant tops only, without the need for N analyses, using statistical analyses. A previously proposed method which employs the linear relation between N-yield and dry matter yield was shown to apply only when data for strains which are not fully effective are excluded.It is postulated that symbiotically fixed N forms a different compound from that resulting from applied mineral N and that this compound cannot be remobilised before flowering.deceased     

A pellet method to demonstrate nitrogen fixation by free-living rhizobia

Summary A method is described to demonstrate nitrogen fixation by free-living Rhizobium cells. After aerobic growth in a nutrient solution, the bacteria are centrifuged. Acetylene reduction by the rhizobial cells in the pellet can be measured within a few days. Hydrogen gas frequently stimulates acetylene reduction.     

Nitrogenase — hydrogenase relationships in Rhizobium japonicum

The conditions necessary for coordinate derepression of nitrogenase and O₂-dependent hydrogenase activities in free-living cultures of Rhizobium japonicum were studied. Carbon sources were screened for their ability to support nitrogenase, and then hydrogenase activities. There was a positive correlation between the level of nitrogenase and corresponding hydrogenase activities among the various carbon substrates. The carbon substrate -ketoglutarate was able to support the highest levels of both nitrogenase and hydrogenase activities. When cells were incubated in -ketoglutarate-containing medium, without added H₂ but in the presence of acetylene (to block H₂ evolution from nitrogenase) significant hydrogenase activity was still observed. Complete inhibition of nitrogenase-dependent H₂ evolution by acetylene was verified by the use of a Hup^- mutant. Hydrogen is therefore not required to induce hydrogenase. The presence of 10% acetylene inhibited derepression of hydrogenase. Constitutive (Hup^c) mutants were isolated which contained up to 9 times the level of hydrogenase acitivity than the wild type in nitrogenase induction medium. These mutants did not have greater nitrogenase activities than the wild type.This is contribution number 1254 from the Department of Biology and the McCollum-Pratt Institute Abbreviations: -Ketoglutarate-containing medium (LOKG) and pre-adaptation medium (SRM) as described in Materials and methods     

Comparison of geographically distant populations of Rhizobium isolated from root nodules of Phaseolus vulgaris

Seventy-two rhizobial strains were isolated from the root nodules of french beans ( Phaseolus vulgaris ). They were sampled from two geographically distant field populations and 18 additional different sites in France. They were characterized by a) plasmid profiles, (b) RFLP analysis of total cellular DNA using various chromosomal and symbiotic gene probes (including nif H from Rhizobium etli bv. phaseoli ) and c) their ability to nodulate a potential alternative host, L. leucocephala. Over half of the isolates were ascribed to Rhizobium leguminosarum bv. phaseoli on the basis of the hybridization analysis, the possession of multiple copies of nif H and their inability to nodulate L. leucocephala. The remaining isolates belonged to 2 groups which were shown to be genomically distinct from R. leguminosarum bv. phaseoli, R. etli bv. phaseoli and R. tropici. Most members of these two groups shared with R. tropici the ability to nodulate L. leucocephala and, for isolates of only one of these groups, the presence of one copy of nif H. Members of each of the 3 taxa were widely distributed in France and circumstantial evidence of pSym transfer between them was shown. R. leguminosarum bv. phaseoli and one of the two novel groups co-occurred within the two geographically distant populations. Individual genotypes were conserved between them. The finding of a third taxon at various other locations indicated additional diversity among rhizobia nodulating beans.     

Identification and characterization of flavonoids in the root exudate of Robinia pseudoacacia

 Eight compounds exuded from young roots of black locust (Robinia pseudoacacia) were separated by two-dimensional HPTLC, by HPLC and GC, and were identified by spectroscopic methods (ultraviolet/visible spectroscopy and mass spectrometry) as 4′,7-dihydroxyflavone, apigenin, naringenin, chrysoeriol and isoliquiritigenin. Structural assignments were confirmed by comparison with authentic standards. The capacity to induce β-galactosidase activity in Rhizobium sp. NGR234 containing a nod box::lacZ fusion on plasmid pA27 identified these flavonoids and the chalcone as nod gene inducers. This indicates the important role of these compounds in nodulation of this legume tree. Received: 26 July 1996 / Accepted: 9 September 1996     

Effect of two granular nematicides on growth and nodulation ofArachis hypogea L.

Summary The effect of two granular nematicidesviz. oxamyl and fenamiphos, on the nodulation and growth of Rhizobium inoculatedArachis hypogaea L. was studied in glasshouse and field trials. In the glasshouse trial at the suggested rates of application shoot fresh weight was significantly reduced by oxamyl whilst root fresh weight was similarly affected by fenamiphos. In the field trial vegetative growth and plant emergence were significantly reduced by both nematicides. Nodulation at the higher rates of application was increased by both oxamyl and fenamiphos whilst oxamyl caused a significant increase in pod number at the highest rate of application.