Abstract: Novel strains of isolated and purified bacteria have been identified which have the ability to degrade petroleum hydrocarbons including a variety of PAHs. Several isolates also exhibit the ability to produce a biosurfactant. The combination of the biosurfactant-producing ability along with the ability to degrade PAHs enhances the efficiency with which PAHs may be degraded. Additionally, the biosurfactant also provides an additional ability to bind heavy metal ions for removal from a soil or aquatic environment.

Abstract: A method for converting organic solid waste into humic products and the corresponding apparatus, in which a preliminary aerobic fermentation of the organic solid waste is performed, followed by a second aerobic fermentation of the organic waste in the presence of specific microorganisms in order to provide a basic precursor on which the selective metabolization occurs of specific bacterial strains to provide the humic product.

Abstract: A method is provided for enhancing oxidation of methyl bromide during agricultural fumigations of fields using a fumigant containing methyl bromide. The method comprises adding a methylotrophic bacterium to the soil in an amount effective to provide bacterial oxidation of the methyl bromide. The bacterium preferably comprises a bacterium isolated from agricultural soil, and, in a specific embodiment, comprises a 16S ribosomal RNA gene sequence in the Alpha subgroup of Proteobacteria designated strain IMB-1 (ATCC 202197). The fumigant also includes chloropicrin in an amount reduced to a level which permits said bacterial oxidation but while still enables the chloropicrin to serve as a warning agent for excessive release of methyl bromide from the soil. The soil can be pretreated with methyl iodide. The bacterium is applied to the soil as freeze-dried bacterial cells during the fumigation operation.

Abstract: A test kit and method for the highly sensitive detection of specific analytes in a sample is provided. The presence of the analyte in the sample results in a decrease in the concentration of a growth inhibiting substance leading to proliferation of cells in the region of the analyte. The presence or absence of the analyte is determined by detecting the presence of increased numbers of cells. Assay sensitivity is accounted for by the exponential amplification of cell number that occurs during cell proliferation in the presence of analyte.

Abstract: A microbiological assay for chemicals, which uses a cell and a reducing dye to quantitatively measure inhibition of electron transport in the cell membrane as a function of chemicals in the substance being tested, is disclosed. This assay and method is reliable, simple, fast, and inexpensive, requires a minimum amount of durable equipment, and avoids the need for the use of live animals as the indicator organisms. The assay is particularly useful for testing for toxicity in food products, environmental, medical and industrial processes, sewage treatment, effluent, agricultural wastes, and chemical dumps.

Abstract: Microorganisms of interest are capable of utilizing .alpha.-imino carboxamides, in the form of the racemate or of its optically active isomers, of the general formula ##STR1## wherein A together with --NH-- and --CH-- is an optionally substituted 5- or 6-membered saturated heterocyclic ring, as sole nitrogen source, and converting (RS)-.alpha.-imino carboximides of Formula I into an S-.alpha.-amino carboxylic acid of the general formula ##STR2## These microorganisms are useful also for biconversion of an (RS)-.alpha.-imino carboxamide of Formula I into an S-.alpha.-amino carboxylic acid.

Abstract: Methods and compositions for increasing the availability of soluble phosphate and fixed nitrogen for legume:Rhizobium symbioses involving co-inoculating legume seeds with a phosphate-solubilizing soil fungus, Penicillium bilaii, and Rhizobium spp. prior to planting. The two types of microorganisms do not appear to compete for nutrients in the rhizosphere of legumes and the P. bilaii provides a source of available phosphorus for use by the plant without adversely affecting the nitrogen fixation ability of the Rhizobium spp.; and indeed such ability is enhanced. The invention is used to increase the efficiencies of nodulation, nitrogen fixation and legume crop production.

Abstract: An essentially pure substance has a structure of a Nod factor or one of its analogues. The Nod factor is characterized by the fact that its biosynthesis is controlled by at least one nodulation gene (nodA,B,C) common to the Rhizobiaceae, in particular to the genera Rhizobium, Bradyrhizobium, Sinorhizobium and Azorhizobium. This substance consists of a lipo-oligosaccharide which is not a derivative of the exopolysaccharides and which has the general formula (I). In formula (I), the Nod factor of which it has the structure is a plant-specific symbiotic signal and is capable of enhancing the capacity of the bacteria of infect the host plant with which it is associated and/or of accelerating the formation of nodules on the host plant with which it is associated and/or of inducing the transcription of symbiotic genes of the leguminoseae. Applications to the treatment of plants and as an active therapeutic agent in humans and animals.

Abstract: There is provided an economical, efficient and industrially useful method of producing vitamin B.sub.12 which comprises cultivating a microorganism belonging to Rhizobium cobalaminogenum FERM BP-4429 which is capable of producing vitamin B.sub.12 in a culture medium and recovering the vitamin B.sub.12. The microorganism is isolated from soil and has no formation of acids from L-arabinose. Further, there are no straight chain fatty acids 21:1 or 19:0.

Abstract: There is provided an economical, efficient and industrially useful method of producing vitamin B.sub.12 which comprises cultivating a microorganism belonging to Rhizobium cobalaminogenum which is capable of producing vitamin B.sub.12 in a culture medium to produce vitamin B.sub.12 and recovering the produced vitamin B.sub.12. There are also provided a microorganism belonging to Rhizobium cobalaminogenum which is capable of producing vitamin B.sub.12 and a microorganism Rhizobium cobalaminogenum 27B74 (FERM BP-4429).

Abstract: The invention includes a consensus nucleotide sequence found in the promoter regions of those Rhizobium and Bradyrhizobium nodulation genes which are activated by chemical inducer(s) in legume exudate. A promoter comprising the consensus sequence is a legume exudate-inducible promoter. A recombinant gene comprising a structural gene and a legume exudate-inducible promoter is selectively expressed in the presence of legume exudate or chemical inducer(s) and a functional nodD gene. The consensus nucleotide sequence isATCCAYNNYGYRGATGNWYKYKATCSAAWCAATCRATTTTACCARWYYKNSRR whereN is A, G, C or TY is C or TR is A or GW is A or TK is G or T andS is C or G.

Abstract: Methods for the isolation of mutant rhizobial strains with improved competition of nodulation (Comp.sup.+) is presented. Mutants are selected for constitutive expression of inducible nod genes (in the absence of inducer) and screened for hyperinduction of nod genes in the presence of inducer. Mutants which are defective in nodulation or in symbiotic nitrogen fixation are eliminated from further testing. Selections for constitutive nod expression and for hyperinduction of nod genes are facilitated by the use of plasmid carrying a selectable marker fused to an inducible nod gene and a plasmid carrying a reporter gene fused downstream of an inducible nod gene. The methods exemplified are particularly useful for the isolation and identification of Comp.sup.+ mutants of Bradyrhizobium japonicum.

Abstract: A bacterial inoculum including a rhizospheric bacterium with increased dicarboxylic acid membrane permease activity.

Abstract: There is disclosed a process for preparing optically active 3-phenylglycidic acid ester compound, which comprises permitting a culture broth, cells or treated cells of a microorganism having an ability of stereoselectively hydrolyzing a (2R, 3S)-3-phenylglycidic acid ester compound to act on a racemic 3-phenylglycidic acid ester compound which may also have a substituent on the phenyl group, thereby hydrolyzing the (2R, 3S) optically active isomer and separating and collecting the (2S, 3R) antipode from the reaction mixture.

Abstract: A novel approach for the increased dinitrogen fixation of leguminous plants wherein the root nodulation of the leguminous plant is controlled so as to promote nodulation by a desired bradyrhizobial strain and to inhibit nodulation of an undesired bradyrhizobial strain. Novel leguminous plant lines are provided which restrict nodulation by undesired bradyrhizobial strains while allowing nodulation of a desired bradyrhizobial strain. The plant lines are produced by conventional breeding. Recombinant bradyrhizobial strains are also provided which are characterized by a propensity for root nodulation of a predetermined leguminous plant. The recombinant bradyrhizobial strains are produced by genetic engineering.

Abstract: The chemical compounds that induce expression of nodulation genes of Bradyrhizobium japonicum strains have been identified. The isoflavones daidzein and genistein were identified as soybean exudate components that induce nod-genes. Other isoflavones, including 7-hydroxyisoflavone, 5,7-dihydroxyisoflavone, biochanin A, formononetin and prunetin, were also found to act as inducers. In addition, several flavonoid compounds including 4',7-dihydroxyflavone, apigenin, kaempferol and coumestrol, were also found to act as inducers. Nodulation inducing compositions and a method for selectively activating the expression of a gene under the control of a soybean exudate inducible promoter in B. japonicum in response to these inducer molecules are described. In addition, soybean inoculating compositions containing the nod-gene inducers and inoculating methods employing these compositions are described.

Abstract: Rhizobia transformants that nodulate and fix nitrogen in non-legumes. The nodulated non-legume plants can be grown without nitrogenous fertilizer and have at least the same or higher protein content, dry matter content and nitrogen content than their non-nodulated counterparts which are fertilized by the addition of nitrogenous fertilizer. The straw remaining after harvesting the nodulated non-legumes is also high in protein content.

Abstract: The sequences of a Rhizobium bacteria responsible for competitiveness with respect to plant nodulation have been isolated and permanently transferred to superior nodulating Rhizobium genome. This has resulted in a stable construct that can form a plant inoculant that yields effective nodulation, while reducing the risk of suppression by other bacteria in the environment.

Abstract: The nodulation ability and/or competition characteristics of strains of Rhizobium japonicum is enhanced by incubating the strains in a nodulation and/or competitive enhancing effective amount of one or more soybean lectins, or in composition comprising a nodulation and/or competitiveness enhancing effective amount of such lectins such as soybean seed exudate. An inoculum can be prepared containing a carrier, Rhizobium japonicum and lectin.

Abstract: E. coli mutants which are capable of expressing cloned proteins in a highly stable manner are described.

Abstract: Nodulation gene-inducing factors have been isolated and identified. A number of substituted flavones, some of which are components of exudates and extracts of leguminous plants, have been found to induce expression of certain nodulation genes in Rhizobium strains. Three components of clover extract 7,4'-dihydroxyflavone, 7,4'-dihydroxy-3'methoxyflavone, and 4'-hydroxy-7-methoxyflavone have been identified as nodulation inducers of certain Rhizobium trifolii nod genes. A number of other flavonoids including flavones, flavanones, flavanols and dihydroflavanols, not necessarily found in legume exudates, were also found to induce expression of certain nodulation genes.

Abstract: This invention provides useful promoters from the R. trifolii nifH gene for the construction of recombinant molecules to regulate foreign genes for expression under desired conditions. In particular, the promoters act to control expression of the foreign genes within root nodules formed by rhizobial bacterial strains in symbiotic combination with host plants.A rhizobium diagnostic segment (RDS) is also provided comprising a DNA segment found at more than one location in rhizobia, the RDS being species-specific, and detectable by DNA hybridization under stringent conditions. A recombinant plasmid comprising a RDS and a bacterial strain containing the plasmid are provided. Methods are provided for identifying species and strains of field isolates of Rhizobium, using RDS's. One RDS exemplified comprises 5' sequences from the R. trifolii nifH gene.

Abstract: The type II restriction endonuclease RleAI has the following recognition sequence: ##STR1## cleaves DNA at the cleavage site indicated by the arrows, and is preferably obtainable from microorganisms of the genus Rhizobium. It can be used to recognize and cleave the double-stranded DNA sequence CCCACA(N).sub.12/9 and its complementary sequence.

Abstract: Plant protoplasts are exposed for interaction with an external agent by exposing the plasma membrane of protoplasts in the root hairs of the plant without release of protoplasts from the plant. This may be achieved by enzymatically degrading the plant cell wall at the apices of root hairs under conditions which expose the plasma membrane whilst maintaining or readily permitting restoration of the essential functional integrity of the plant. Enzymatic degradation of the cell wall of the apices of root hairs is possible in a wide range of crop species and other species and to expose the plasma membrane with partial protoplast release. This enables direct interaction with plasmids, viruses and micro-organisms at the surface of the plasma membrane of their exposed protoplasts, or the fusion of these with isolated protoplasts of other plant systems.

Abstract: A method of transforming eukaryotic or prokaryotic hosts sensitive to an antibiotic of the phleomycin family to confer resistance to the antibiotic is disclosed in which a phleomycin resistance gene is used as a selectable marker.

Abstract: Novel constructs are provided containing DNA fragments comprising a Rhizobium nodulation gene divergent transcriptional initiation regulatory region. The region is responsive to plant exudate in the presence of a nod D gene product. When associated with plants, the region can control the expression of structural genes, such as agents active in protecting plants or inducing their growth.This invention was funded at least in-part by a grant from the National Institutes of Health. The U.S. government may have certain rights in this invention.

Abstract: A method of controlling expression of a DNA segment under the control of a nod gene promoter which comprises administering to a host containing a nod gene promoter an amount sufficient to control expression of the DNA segment of a compound of the formula: ##STR1## in which each R is independently H or OH, is described.

Abstract: The isolation and characterization of nodD-related genes in soybean nodulating Rhizobium japonicum is described. In R. japonicum USDA 191 two such genes have been identified, which although related in structure, have different functional properties. These nodD genes are functionally distinct from each other and from those nodD genes of other strains of Rhizobium that have been isolated and characterized to date. In particular, nodD-r1 has been found to affect nodulation on soybean and to be associated with exopolysaccharide production. In contrast, nodD-r2 affects nodulation on the tropical legume siratro. The coding sequences of both nodD genes are provided. The promoter regions of the nodD genes have also been isolated. The genes described herein, including both structural genes and their promoter regions, can be used in combination with other genetic constructs to enhance the efficiency and competitiveness of nodulation.

Abstract: Rhizobial ferredoxin genes and proteins are provided. Rhizobial ferredoxins are useful to enhance the nitrogenase systems of rhizobia. Useful rhizobial ferredoxin diagnostic segments are also provided comprising DNA sequences encoding the characteristic ferredoxin cysteine residue patterns. Exemplied rhizobial ferredoxin genes are fixX of Rhizobium trifolii and Rhizobium meliloti Between fixC and nifA, fixY of Rhizobium meliloti down stream from nifB, and frxA of Bradyrhizobium japonicum.

Abstract: A method of increasing nodulation of a plant capable of being nodulated by Bradyrhizobium sp. (Parasponis) is provided comprising infecting said plant with a Bradyrhizobium sp. (Parasponia) species mutated such that nodK is non-functional. One such method of mutation involves insertion upstream of nodABC constitutive promoter sequences capable of activating nodABC. The invention is exemplified by the inoculation of siratro with mutated Bradyrhizobium sp. (Parasponia) mutated by insertion of nptII promoter sequences in nodK upstream of nodABC so as to activate nodABC. As compared with inoculation with the corresponding wild-type Bradyrhizobium sp. (Parasponia), nodulation onset occurred five days earlier, nodulation number was doubled, and a 120% enhancement of plant yield was shown. Similar results were obtained regardless of the orientation of the nptII promoter-containing sequences within the nodK coding sequence.

Abstract: A method is disclosed for producing novel strains of Rhizobium bacteria for legume inoculants which are both competitive and have high nitrogen fixation characteristics. The method involves first isolating and identifying the naturally predominant strains of the Rhizobium species of interest in a particular locale and then mutagenizing that strain to produce mutant strains which retain the competitiveness and add enhanced nitrogen fixation capabilities.

Abstract: Nodulation regulatory genes (nodD genes) of Bradyrhizobium japonicum strains have been isolated and sequenced. Recombinant DNA molecules and vectors containing these regulatory genes are described. These genes, molecules and vectors are useful in the genetic engineering of Rhizobium and Bradyrhizobium strains. A method for selective expression of structural genes in response to the application of chemical factors which induce B. japonicum nod genes which employs these genes is described.

Abstract: A prototrophic revertant of a Bradyrhizobium japonicum tryptophan auxotroph was isolated and found to fix more nitrogen symbiotically than wild-type bacteria. The increase in nitrogen fixation is due to an increase in nodule mass because of an increase in nodule number. The physiological basis for this improved symbiosis appears to be an alteration of the tryptophan biosynthetic pathway.

Abstract: A method of transforming eukaryotic or prokaryotic hosts sensitive to an antibiotic of the phleomycin family to confer resistance to the antibiotic is disclosed in which a phleomycin resistance gene is used as a selectable marker.

Abstract: Recombinant DNA clones according to the invention contain nodABCDIJL genes of B. japonicum. These clones can be used to practice a method wherein a nod- strain of Rhizobium or Bradyrhizobium bacteria lacking one or more of these genes is converted to a nod+ strain by transfering one or more of the missing genes to the strain. According to a further aspect of the invention, nucleotide sequences have been identified which control the expression of nod LABC genes of Bradyrhizobium japonicum in the presence of flavones produced by the plant roots. By linking such sequences with a gene having a desired characteristic, such as a gene encoding for production of a plant parasite toxin, bacterial can be created which selectively express the characteristic only in the presence of the flavone which triggers expression of the gene.

Abstract: Recombinant DNA clones according to the invention contain a host range gene of B. japonicum which is contained within a 3.3 kb HindIII restriction fragment from the B. japonicum chromosome. Mutations to this gene change the ability of the bacteria to nodulate different plants. In one example, a Tn5 insertion to this region caused the strain to lose its ability to nodulate siratro, but nodulation of soybeans was unaffected. This gene may be manipulated to extend or restrict the host range of the strain for agricultural purposes.

Abstract: The present invention comprises a biologically pure culture of a cellulose-producing microorganism, preferably a prokaryote. This cellulose-producing microorganism is capable, during fermentation in an aqueous nutrient medium containing assimilable sources of carbon, nitrogen and inorganic substances, of reversal of direction of cellulose ribbon extrusion. This reversal of direction of cellulose ribbon extrusion results on the cellulose-producing microorganism shuttling, at least periodically, first in one direction and then in the other direction along a length of an earlier-deposited cellulose ribbon to add another cellulose ribbon thereto and produce a cellulose ribbon-bundle having a width of at least two cellulose ribbons.The cellulose-producing microorganism of the present invention may be of the genus Acetobacter, Agrobacterium, Rhizobium, Pseudomonas or Alcaligenes, preferably of the genus Acetobacter and more preferably of the species Acetobacter xylinum or Acetobacter pasteurianus.

Abstract: Mutant strains of Bradyrhizobium japonicum having enhanced nodulation properties were created by transposon mutagenesis of known Bradyrhizobium japonicum strain I-110. The mutant strains grow well in a yeast-extract mannitol medium, produce extracellular polysaccharides at a level greater than the parent strain under appropriate conditions, are capable of growth on a nutirent medium containing a normally inhibitory amount of succinic acid, and contain a 21 Kdalton protein absent from the parent strain. Such strains can be used to inoculate soil in which soybean plants are grown, resulting in improved plant yields.

Abstract: A symbiosis plasmid from a fast-growing Rhizobium japonicum donor strain can be transferred to Rhizobium recipient strains, the recipient strains being previously incapable of forming an effective symbiotic relationship with plants of certain Glycine (e.g. soybean) varieties. The recipient strains harboring the symbiosis plasmid will form effective symbioses with soybean plants of said certain varieties, the transferred symbiosis plasmid conferring the host range or specificity of the donor strain on the recipient strains. Methods, strains, and plasmids useful for practice of this invention are also provided.

Abstract: Inocula having a long storage life and improved resistance to temperature and rehydration and a method of preparing them are disclosed. The method of the invention comprises admixing a culture medium containing microorganisms of the genus Rhizobium and at least one carbohydrate source in a polymer gel and then lowering the water activity of the resulting inoculum and maintaining the water activity at less than 0.1.

Abstract: Nodulation-enhancing mutants of Rhizobium species are prepared by growing a wild-type strain in a medium as the only carbon source at a sub-minimum concentration for normal growth, the mutant cells thus obtained are further selected by comparative nodulation to obtain mutants which produce two to three times as many nodules as the corresponding wild-type strain.

Abstract: A Tn-Mob element is provided comprising a transposon with a Mob-site inserted therein. Vectors and bacterial strains are provided containing the Tn-Mob element, as well as methods for its use. Such methods include the production of plasmid free bacterial strains.

Abstract: Novel strains of Rhizobium phaseoli which do not produce acidic extracellular polysaccharides or their constituents, which produces cyclic .beta.-1,2-glucan effectively.

Abstract: Bacteria of the genus Rhizobium nodulate legumes and are responsible for nitrogen fixation. The energetics of this process are improved if hydrogen uptake ability (Hup) is imparted to the bacteria to "recycle" some of the hydrogen lost during the nitrogen fixation. Such Hup is observed in a naturally occurring strain of Rhizobium leguminosarum but it was not known how to transfer it to other strains. The present invention provides recombinant plasmids pIJ1008 and pIJ1007 which enable it to be transferred. These plasmids are formed from a transmissible plasmid, synthesized to include a drug-resistance marker, and a plasmid of the naturally occurring strain. Cultures containing the plasmids have been deposited in a culture collection and can be used, after appropriate further crosses as described, to impart Hup to other strains of Rhizobium.

Abstract: Novel processes are provided for the embedding of microorganisms within a polymer matrix wherein the matrix comprises a polymer gel having a base of at least one polymer selected from the group of polysaccharides, in which said polymer is at least partially cross-linked. The embedded microorganism products are useful in the inoculation of leguminous and non-leguminous plants in order to increase their nitrogen-fixing potential and nutrition, respectively.

Abstract: Viscous water for use in oil extraction is produced continuously by bacterial action. A carbohydrate substrate, nitrogen-containing mineral salts and growth factors are introduced into a first fermenter chamber containing bacteria of the genus Rhizobium capable of excreting polysaccharides having viscosity increasing properties and aqueous product, from which the bacteria are separated, is collected from the outlet of this chamber and recycled to the inlet of chamber. Into a second viscosifier chamber there is introduced carbohydrate substrate for synthesis of polysaccharide and bacteria recovered from the first chamber in an amount approximately equal to that which is removed with the viscous product from the second chamber and there is collected from the outlet of the second chamber the viscous product at a rate approximately equal to that of the carbohydrate substrate at the inlet in order to maintain equilibrium in the system.

Abstract: This invention relates to Rhizobium strains having good infecting and nitrogen-fixing characteristics and which are resistant to fungicides. These strains are produced by cultivating a Rhizobium strain sensitive to a particular fungicide in the presence of an amount of the fungicide, and for a time less than that sufficient to kill the entire Rhizobium population but sufficient to kill a majority of the population. The remaining Rhizobium population is isolated and recultured in the presence of an increased amount of the fungicide, again an amount and a time less than sufficient to kill the entire Rhizobium population, but sufficient to kill a majority of the population. This procedure is repeated for sufficient passages, with increasing amounts of said fungicide, to provide a Rhizobium strain sufficiently resistant to said fungicide so that the Rhizobium strain multiplies and enters into a nitrogen-fixing symbiosis in the presence of agriculturally effective amounts of the fungicide.

Abstract: A method of packaging, and the package so produced, for maintaining living organisms viable for a long period of time. The living organisms, such as bacteria, fungi, algae, etc., are mixed with a carrier, such as peat. The organism-carrier mixture is then disposed in a package, such as a heat-sealable plastic envelope, and a gaseous atmosphere is provided in the package effective to induce and maintain substantial nonvegatative state formation of the organisms. Some organisms will form cysts, others spores, but whatever nonvegetative state is assumed, the organisms will be much less susceptible to heat, cold, starvation, and other adverse environmental factors. Suitable gaseous atmospheres include nitrogen, helium, and argon gases. The package is then sealed to prevent contamination of the atmosphere therein.