Bra.dy.rhi.zo' bi.um. Gr. adj.bradusslow; M.L. neut. n.Rhizobiuma bacterial generic name; M.L. neut. n.Bradyrhizobiumthe slow (growing) rhizobium.Proteobacteria / Alphaproteobacteria / Rhizobiales / Bradyrhizobiaceae / BradyrhizobiumRods 0.5–0.9 × 1.2–3.0 µm.Commonly pleomorphic under adverse growth conditions. Usually contain granules of poly‐β‐hydroxybutyrate that are refractile by phase‐contrast microscopy.Nonsporeforming.Gram negative.Motile by one polar or subpolar flagellum. Fimbriae have not been described.Aerobic,possessing a respiratory type of metabolism with oxygen as the terminal electron acceptor.Optimal temperature 25–30°C. Optimal pH, 6–7, although lower optima may be exhibited by strains from acid soils. Colonies are circular, opaque, rarely translucent, white, and convex, and tend to be granular in texture; they do not exceed 1.0 mm in diameter in less than 5–6 days incubation on A1EG medium.Turbidity develops only after 3–4 days in agitated broth.Generation times are 9–18 h. Chemoorganotrophic, utilizing a range of carbohydrates and salts of organic acids as carbon sources, without gas formation; arabinose and other pentoses are preferred carbon sources. Cellulose and starch are not utilized. Produce an alkaline reaction in mineral salts medium containing mannitol and/or many other carbohydrates.Growth on carbohydrate media is usually accompanied by extracellular polysaccharide slime production particularly with glycerol,gluconate,or mannitol. Some strains can grow chemolithotrophically in the presence of H2, CO2, and low levels of O2. Ammonium salts, usually nitrates, and some amino acids, can serve as nitrogen sources. Peptone is poorly utilized (except for strains isolated fromLotononis). Casein and agar are not hydrolyzed. There is usually no requirement for vitamins with the rare exception of biotin, which also may be inhibitory to some strains. 3‐Ketoglycosides are not produced (Bernaerts and De Ley, 1963).The organisms are characteristically able to enter the root hairs of tropical‐zone and some temperate‐zone leguminous plants(family Leguminosae)and incite the production of root nodules,in which the bacteria occur as intracellular nitrogen‐fixing symbionts. Some strains, especiallyB.elkanii, fix nitrogen in the free‐living state when examined under special conditions.The mol%G+C of the DNA is: 61–65.Type species:Bradyrhizobium japonicum(Kirchner 1896) Jordan 1982, 137 (“Rhizobacterium japonicum” Kirchner 1896, 221;Rhizobium japonicum(Kirchner 1896) Buchanan 1926, 90.)

Rhi.zo' bi.um . Gr. n. rhiza a root; Gr. n. bios life; M.L. neut. n. Rhizobium that which lives in a root. Proteobacteria / Alphaproteobacteria / Rhizobiales / Rhizobiaceae / Rhizobium Rods 0 . 5–1 . 0 × 1 . 2–3 . 0 µm . Nonsporeforming. Gram negative. Motile by 1–6 peritrichous flagella . Fimbriae have been described on some strains. Aerobic , possessing a respiratory type of metabolism with oxygen as the terminal electron acceptor. Optimal temperature for growth, 25–30°C; some species can grow at temperatures >40°C. Optimal pH for growth, 6–7; range pH 4–10. Generation times of Rhizobium strains are 1.5–5.0 h. Colonies are usually white or beige, circular, convex, semi‐translucent or opaque, raised and mucilaginous, usually 2–4 mm in diameter within 3–5 days on yeast‐mannitol‐mineral salts agar ( YMA ). Growth on carbohydrate media is usually accompanied by copious amounts of extracellular polysaccharide . Pronounced turbidity develops after 2 or 3 days in aerated or agitated broth. Chemoorganotrophic, utilizing a wide range of carbohydrates and salts of organic acids as sole carbon sources, without gas formation. Cellulose and starch are not utilized. Produce an acidic reaction in mineral‐salts medium containing mannitol or other carbohydrates . Ammonium salts, nitrate, nitrite, and most amino acids can serve as nitrogen sources. Strains of some species will grow in a simple mineral salts medium with vitamin‐free casein hydrolysate as the sole source of both carbon and nitrogen, but strains of many species require one or more growth factors such as biotin, pantothenate, or nicotinic acid. Peptone is poorly utilized. Casein, starch, chitin, and agar are not hydrolyzed. All known Rhizobium species include strains which induce hypertrophisms in plants as root nodules with or without symbiotic nitrogen fixation . Some cells of symbiotic bacterial species enter root hair cells of leguminous plants (Family Leguminosae) via invagination or by wounds (“crack entry”) and elicit the production of root nodules wherein the bacteria engage as intracellular symbionts, usually fixing nitrogen. Many well‐defined nodulation ( nod ) and nitrogen fixation ( nif ) genes are clustered on large plasmids or megaplasmids (pSyms). Plasmid transfer between species results in the expression and stable inheritance of the particular plant‐interactive properties of the plasmid‐donor species. Plant host specificity is usually for a few legume genera but may, in some strains, include a wide variety of legume genera and is to some extent determined by the chemical structure of the lipochito‐oligosaccharide Nod factors produced. These chitin‐like molecules induce nodule organogenesis in the absence of bacteria. In root nodules the bacteria occur as endophytes that exhibit pleomorphic forms , termed “ bacteroids ”, which reduce or fix gaseous atmospheric nitrogen into a combined form utilizable by the host plant . The mol % G + C of the DNA is : 57–66. Type species : Rhizobium leguminosarum (Frank 1879) Frank 1889, 338 ( Schinzia leguminosarum Frank 1879, 397.)

Me.so.rhi.zo' bi.um . Gr. adj. mesos middle; M.L. neut. n. Rhizobium bacterial generic name; M.L. neut. n. Mesorhizobium the meso ‐growing rhizobium, referring to the growth rate intermediate between those of the genera Rhizobium and Bradyrhizobium . Proteobacteria / Alphaproteobacteria / Rhizobiales / Phyllobacteriaceae / Mesorhizobium Rods 0.4–0.9 × 1.2–3.0 μm. Commonly pleomorphic under adverse growth condition or in the root nodules as bacteroids . Usually contain granules of poly‐β‐hydroxybutyrate, which are refractile by phase‐contrast microscopy. Nonsporeforming. Gram negative. Motile by one polar or subpolar flagellum or by peritrichous flagella. Aerobic, possessing a respiratory type of metabolism with oxygen as the terminal electron acceptor. Optimal temperature 25–30°C. Optimal pH 6–8. Growth in yeast extract–mannitol agar (YMA) 1 produces colonies that are circular, convex, semitranslucent, raised, and mucilaginous, 2–4 mm diameter within 5–6 d for some species or 1 mm after a 7‐d incubation for other species. The generation times of Mesorhizobium strains range from 4–15 h . Chemoorganotrophic, utilizing a wide range of carbohydrates and salts of organic acids as carbon sources without gas production. Cellulose and starch are not utilized. Produces an acidic reaction in YMA . Ammonium salts, nitrates, urea, and most amino acids are utilized as nitrogen sources. Peptone is poorly utilized. Mesorhizobium strains are only weakly proteolytic, but can produce a slow digestion in litmus milk; some strains form a clear serum zone. Some strains require thiamin, nicotinamide, and riboflavin for growth. The organisms are characteristically able to invade the root hairs of a wide range of temperate, subtropical, and tropical leguminous plants, inciting production of root nodules where the bacteria reduce atmospheric nitrogen into a combined form available for the host plants . All strains exhibit host specificity. The mol % G + C of the DNA is : 59–64. Type species : Mesorhizobium loti (Jarvis, Pankhurst, and Patel 1982) Jarvis, van Berkum, Chen, Nour, Fernandez, Cleyet‐Marel and Gillis 1997, 898 ( Rhizobium loti Jarvis, Pankhurst, and Patel 1982, 378.)

Rhi.zo.bi.a' les . M.L. n. Rhizobium type genus of the family Rhizobiaceae ; ‐ ales ending to denote an order; M.L. fem. n. Rhizobiales the Rhizobium order. Proteobacteria / Alphaproteobacteria / Rhizobiales

'Ca. Liberibacter asiaticus,' an insect-vectored, obligate intracellular bacterium associated with citrus-greening disease, also called "HLB," is a member of the Rhizobiales along with nitrogen-fixing microsymbionts Sinorhizobium meliloti and Bradyrhizobium japonicum, plant pathogen Agrobacterium tumefaciens and facultative intracellular mammalian pathogen Bartonella henselae. Comparative analyses of their circular chromosomes identified 514 orthologous genes shared among all five species. Shared among all five species are 50 identical blocks of microsyntenous orthologous genes (MOGs), containing a total of 283 genes. While retaining highly conserved genomic blocks of microsynteny, divergent evolution, horizontal gene transfer and niche specialization have disrupted macrosynteny among the five circular chromosomes compared. Highly conserved microsyntenous gene clusters help define the Rhizobiales, an order previously defined by 16S RNA gene similarity and herein represented by the three families: Bartonellaceae, Bradyrhizobiaceae and Rhizobiaceae. Genes without orthologs in the other four species help define individual species. The circular chromosomes of each of the five Rhizobiales species examined had genes lacking orthologs in the other four species. For example, 63 proteins are encoded by genes of 'Ca. Liberibacter asiaticus' not shared with other members of the Rhizobiales. Of these 63 proteins, 17 have predicted functions related to DNA replication or RNA transcription, and some of these may have roles related to low genomic GC content. An additional 17 proteins have predicted functions relevant to cellular processes, particularly modifications of the cell surface. Seventeen unshared proteins have specific metabolic functions including a pathway to synthesize cholesterol encoded by a seven-gene operon. The remaining 12 proteins encoded by 'Ca. Liberibacter asiaticus' genes not shared with other Rhizobiales are of bacteriophage origin. 'Ca. Liberibacter asiaticus' shares 11 genes with only Sinorhizobium meliloti and 12 genes are shared with only Bartonella henselae.

Sinorhizobium meliloti strain 1021, a nitrogen-fixing, root-nodulating bacterial microsymbiont of alfalfa, has a 3.5 Mbp circular chromosome and two megaplasmids including 1.3 Mbp pSymA carrying nonessential 'accessory' genes for nitrogen fixation (nif), nodulation and host specificity (nod). A related bacterium, psyllid-vectored 'Ca. Liberibacter asiaticus,' is an obligate phytopathogen with a reduced genome that was previously analyzed for genes orthologous to genes on the S. meliloti circular chromosome. In general, proteins encoded by pSymA genes are more similar in sequence alignment to those encoded by S. meliloti chromosomal orthologs than to orthologous proteins encoded by genes carried on the 'Ca. Liberibacter asiaticus' genome. Only two 'Ca. Liberibacter asiaticus' proteins were identified as having orthologous proteins encoded on pSymA but not also encoded on the chromosome of S. meliloti. These two orthologous gene pairs encode a Na(+)/K+ antiporter (shared with intracellular pathogens of the family Bartonellacea) and a Co++, Zn++ and Cd++ cation efflux protein that is shared with the phytopathogen Agrobacterium. Another shared protein, a redox-regulated K+ efflux pump may regulate cytoplasmic pH and homeostasis. The pSymA and 'Ca. Liberibacter asiaticus' orthologs of the latter protein are more highly similar in amino acid alignment compared with the alignment of the pSymA-encoded protein with its S. meliloti chromosomal homolog. About 182 pSymA encoded proteins have sequence similarity (≤ E-10) with 'Ca. Liberibacter asiaticus' proteins, often present as multiple orthologs of single 'Ca. Liberibacter asiaticus' proteins. These proteins are involved with amino acid uptake, cell surface structure, chaperonins, electron transport, export of bioactive molecules, cellular homeostasis, regulation of gene expression, signal transduction and synthesis of amino acids and metabolic cofactors. The presence of multiple orthologs defies mutational analysis and is consistent with the hypothesis that these proteins may be of particular importance in host/microbe interaction and their duplication likely facilitates their ongoing evolution.

MAP3Kα, a gene that encodes a key conserved protein kinase, is responsible for initiating a rapid cascade of cellular events leading to localized cell death. Hypersensitive response, as it is termed, enables genetically resistant plants to limit microbial invasion under the right environmental conditions. Since knowledge of close physically linked genes is important for genome analysis and possibly for improving disease resistance, systematic DNA sequence analysis, gene annotation, and protein BLASTs were performed to identify and characterize genes in close physical proximity to a MAP3Kα-like gene in Beta vulgaris L. US H20. On the same 125 Kb BAC, callose synthase (BvCS) and phytochrome A (PhyA) genes were within 50 Kb of MAP3Kα. The close physical linkage of these genes may result from selection for coordinated responses to disease pressure. Bert, a new chromodomain-carrying gypsy-like LTR retrotransposon, resides within an intron of the BvCS gene, where it is transcribed from the opposing strand.

An intracellular plant pathogen 'Candidatus Liberibacter asiaticus,' a member of the Rhizobiales, is related to Sinorhizobium meliloti, Bradyrhizobium japonicum, nitrogen fixing endosymbionts, Agrobacterium tumefaciens, a plant pathogen, and Bartonella henselae, an intracellular mammalian pathogen. Whole chromosome comparisons identified at least 50 clusters of conserved orthologous genes found on the chromosomes of all five metabolically diverse species. The intracellular pathogens 'Ca. Liberibacter asiaticus' and Bartonella henselae have genomes drastically reduced in gene content and size as well as a relatively low content of guanine and cytosine. Codon and amino acid preferences that emphasize low guanosine and cytosine usage are globally employed in these genomes, including within regions of microsynteny and within signature sequences of orthologous proteins. The length of orthologous proteins is generally conserved, but not their isoelectric points, consistent with extensive amino acid substitutions to accommodate selection for low GC content. The 'Ca. Liberibacter asiaticus' genome apparently has all of the genes required for DNA replication present in Sinorhizobium meliloti except it has only two, rather than three RNaseH genes. The gene set required for DNA repair has only one rather than ten DNA ligases found in Sinorhizobium meliloti, and the DNA PolI of 'Ca. Liberibacter asiaticus' lacks domains needed for excision repair. Thus the ability of 'Ca. Liberibacter asiaticus' to repair mutations in its genome may be impaired. Both 'Ca. Liberibacter asiaticus and Bartonella henselae lack enzymes needed for the metabolism of purines and pyrimidines, which must therefore be obtained from the host. The 'Ca. Liberibacter asiaticus' genome also has a greatly reduced set of sigma factors used to control transcription, and lacks sigma factors 24, 28 and 38. The 'Ca. Liberibacter asiaticus' genome has all of the hallmarks of a reduced genome of a pathogen adapted to an intracellular lifestyle.

A nest of long terminal repeat (LTR) retrotransposons (RTRs), discovered by LTR_STRUC analysis, is near core genes encoding the NPR1 disease resistance-activating factor and a heat-shock-factor-(HSF-) like protein in sugarbeet hybrid US H20. SCHULTE, a 10 833 bp LTR retrotransposon, with 1372 bp LTRs that are 0.7% divergent, has two ORFs with unexpected introns but encoding a reverse transcriptase with rve and Rvt2 domains similar to Ty1/copia-type retrotransposons and a hypothetical protein. SCHULTE produced significant nucleotide BLAST alignments with repeat DNA elements from all four families of plants represented in the TIGR plant repeat database (PRD); the best nucleotide sequence alignment was to ToRTL1 in Lycopersicon esculentum. A second sugarbeet LTR retrotransposon, SCHMIDT, 11 565 bp in length, has 2561 bp LTRs that share 100% identity with each other and share 98-99% nucleotide sequence identity over 10% of their length with DRVs, a family of highly repetitive, relatively small DNA sequences that are widely dispersed over the sugarbeet genome. SCHMIDT encodes a complete gypsy-like polyprotein in a single ORF. Analysis using LTR_STRUC of an in silico deletion of both of the above two LTR retrotransposons found that SCHULTE and SCHMIDT had inserted within an older LTR retrotransposon, resulting in a nest that is only about 10 Kb upstream of NPR1 in sugarbeet hybrid US H20.

McGrath et al., (2004) constructed a Bacterial Artificial Chromosome (BAC) genomic library of sugar beet hybrid H20. In collaboration with Dr. McGrath at MSU, our laboratory successfully identified, sequenced and annotated about 39 Kb (Kuykendall et al., 2007) of a BAC clone carrying NPR1, a key gene controlling disease resistance in plants. The NPR1 protein is required for induced systemic resistance in sugar beet in much the same manner as in Arabidopsis thaliana (Bargabus-Larson and Jacobsen, 2007). Evolutionary conservation of Beta vulgaris FLOWERING LOCUS C that mediates vernalization response was reported (Reeves et al., 2007). Complete gene content of the entire 130-Kb NPR1-carrying BAC was reported (Kuykendall et al., 2008). Functions of predicted protein products of four core plant genes were predicted and conserved microsynteny established in B. vulgaris, Medicago truncatula and Populus trichocarpa, except in M. truncatula where HSF is lacking (Kuykendall et al., 2008). Coe1, a DNA transposase gene within a LTR-retrotransposon, a novel arrangement, was discovered upstream from CaMP on the NPR1 BAC (Kuykendall et. al., 2008). A nest of long terminal repeat LTR-retrotransposons, just upstream of the NPR1 gene, contains both a copia-like and a gypsy-like transposon, SCHULTE and SCHMIDT respectively, within a much older retroelement (Kuykendall et al., in press). In this study, Solanum lycopersicum and Vitis vinifera were assessed for microcolinearity of HSF, NPR1, CaMP and CK1PK genes. HSF, identified by BLAST as a developmental heat shock factor-like protein, and NPR1 proteins activate expression of genes whose products are required for either developmental processes or effective defense against pathogen attack, respectively. CK1 activates expression of genes whose products are involved with chromosome partitioning and circadian rhythm. A role of CaMP protein encoded by CaMP is as yet undefined, but results of experiments herein reported demonstrate that transcription of CaMP in B. vulgaris is upregulated by Erwinia betavasculorum challenge, and it can therefore be inferred that the CaMP protein plays a role in an effective pathogen response.

An understanding of plant molecular genetics underlying programmed responses to pathogens is sought in order to more fully realize the genome’s potential to resist microbial pathogens A previously devised hypothesis was that a cluster of orthologous genes (COG) consisting of four core plant genes in B. vulgaris as well as several other eudicots may contribute to coordinated expression of two or more genes whose products are critical for managing biotic and other environmental stresses. One of these genes, CaMP encodes a 525-amino acid protein product with an N-terminal signal, an IQ domain from amino acid positions 131 to 153, four regions of intrinsic disorder and four phosphorylation sites that roughly correlate with the regions of intrinsic disorder. The most likely calmodulin-binding site on was predicted by Calmodulin Target Database at amino acid positions 201-209. Sugarbeet plants under biotic stress, i.e., experimental Erwinia betavasculorum infection, showed enrichment of a higher level of CaMP-specific RNA transcript compared with un-inoculated control plants. The results of this study suggest that CaMP encodes a chloroplast

Spontaneous azide resistant mutants were isolated from the WT strains of Sinorhizobium fredii, Mesorhizobium ewer, and Rhizobium leguminosarum by trifolii and their symbiotic effectivity was tested on respective host plants either in large test tubes or in Leonard jars A total of 12 mutants were Isolated from the parent strain 54D of Sinorhizobium fredii, 11 mutants from parent strain USDA3383 of Mesorhizobium ewer, and 38 from parent strain ARC100 of Rhizobium leguminosarum by trifolii The host plants Inoculated with a few of the mutants of all the three species of rhizobia resistant to low doses of azide had significantly higher shoot dry weight and ARA in the nodules than the plants inoculated with the parent strains

NPR1 is a gene of central importance in enabling plants to resist microbial attack. Therefore, knowledge of nearby genes is important for genome analysis and possibly for improving disease resistance. In this study, systematic DNA sequence analysis, gene annotation, and protein BLASTs were performed to determine genes near the NPR1 gene in Beta vulgaris L., Medicago truncatula Gaertn, and Populus trichocarpa Torr. & Gray, and to access predicted function. Microsynteny was discovered for NPR1 with genes CaMP, encoding a chloroplast-targeted signal calmodulin-binding protein, and CK1PK, a CK1-class protein kinase. Conserved microsynteny of NPR1, CaMP, and CK1PK in three diverse species of eudicots suggests maintenance during evolution by positive selection for close proximity. Perhaps close physical linkage contributes to coordinated expression of these particular genes that may control critically important processes including nuclear events and signal transduction.

Erwinia carotovora subspecies betavasculorum , also known as E. betavasculorum and Pectobacterium betavasculorum , is a soil bacterium that has the capacity to cause root rot necrosis of sugarbeets . The qualitatively different pathogenicity exhibited by the virulent E. carotovora strain and two avirulent strains, a Citrobacter sp. and an Enterobacter cloacae , was examined using digital analysis of photographic evidence of necrosis as well as for carbohydrate, ethane, and ethylene release compared with uninoculated potato tuber slices. Visual scoring of necrosis was superior to digital analysis of photographs. The release of carbohydrates and ethane from potato tuber slices inoculated with the soft rot necrosis-causing Erwinia was significantly greater than that of potato tuber slices that had not been inoculated or that had been inoculated with the nonpathogenic E. cloacae and Citrobacter sp. strains. Interestingly, ethylene production from potato slices left uninoculated or inoculated with the nonpathogenic Citrobacter strain was 5- to 10-fold higher than with potato slices inoculated with the pathogenic Erwinia strain. These findings suggest that (1) carbohydrate release might be a useful measure of the degree of pathogenesis, or relative virulence; and that (2) bacterial suppression of ethylene formation may be a critical step in root rot disease formation.