Reasons are advanced for removal of Rhizobium ciceri, Rhizobium huakuii, Rhizobium loti, Rhizobium mediterraneum, and Rhizobium tianshanense from the genus Rhizobium and for establishment of Mesorhizobium gen. nov. for these species. A description of the genus Mesorhizobium and amended descriptions of Mesorhizobium ciceri, Mesorhizobium huakuii, Mesorhizobium loti, Mesorhizobium mediterraneum, and Mezorhizobium tianshanense are provided.In a review of root nodule symbioses by Vincent (22), the fast-growing rhizobia associated with Lotus comiculatus and Lupinus densiflorus were recognized as a separate group which merited a specific designation, and the name Rhizobium loti was tentatively proposed for it. Approval for the designation of R. loti as a new species was also voiced at a roundtable discussion on Rhizobium taxonomy associated with the 4th International Congress on Nitrogen Fixation at Canberra, Australia, in 1980. The subsequent publication of the new species in 1982 (9) enabled it to be included in a revised taxonomy of the Rhizobiaceae presented by Jordan in Bergey's Manual of Systematic Bacteriology (12).At that time R. loti was distinguished from other fast-growing rhizobia on the basis of flagellation (l), esterase patterns (14), response to isoflavonoids (18), plant nodulation (11, 12), internal antigens (23), electrophoresis of soluble cellular proteins (19,20), and DNA relatedness (4, 8). More recently, cellular fatty acid analysis was used to reveal differences, useful for identification purposes, between strains of R. loti and strains of the genus Agrobacterium and other Rhizobium or Sinorhizobium species (10). All of these methods differentiate species but give little indication of the relationships between species and genera.A n early indication of the relationship among R. loti, other Rhizobium species, and Sinorhizobium and Agrobacterium species was obtained by studying the intergeneric similarities of rRNA cistrons (7). The results of this analysis indicated that Rhizobium leguminosarum, Rhizobium galegae, Sinorhizobium meliloti, Sinorhizobium fredii, Agrobacterium tumefaciens (biovar l), and Agrobacterium rhizogenes (biovar 2) were all more closely related to one another than they were to R. loti. Subsequent analyses of 16s rRNA gene sequences of species in these genera have confirmed, refined, and extended this observation (21,24,25); the levels of 16s ribosomal DNA sequence similarity between R. loti and other Rhizobium and Agrobacterium species are around 93.5%. Consequently, there has been considerable support for the establishment of a separate genus for R. loti and related root nodule bacteria (5, 13, 27).The original description of R. loti (9) clearly indicated that fast-growing Lotus rhizobia formed part of an extensive plant cross-inoculation group involving plant species in the genera Lupinus, Ornithopus, Lotus, Anthyllis, Caragena, Astragalus, Ononis, Genista, and Mimosa. Jarvis et al. also indicated that the fast-growing Lotus rhizobia were related to rhizobia obtained from s...

A total of 33 strains of fast-growing soybean rhizobia isolated from soil and soybean nodules collected in China and 25 strains belonging to the genera Rhizobium, Bradyrhizobium, and Agrobacteriurn were compared by numerical taxonomic techniques, using 240 different characters, Our results indicated that all of the strains of fast-growing soybean rhizobia which we examined are closely related (guanine-plus-cytosine content, 59.9 to 63.8 mol%) and are separated from Rhizobium and Bradyrhizobium at the generic level. Based on numerical taxonomy, deoxyribonucleic acid (DNA) base ratio determinations, DNA-DNA hybridization data, serological analysis data, the composition of extracellular gum, bacteriophage typing data, and soluble protein patterns, we propose that the fast-growing soybean rhizobia represent members of a new genus rather than a species of Rhizobium (Rhizobium fredii); we propose Sinorhizobium gen. nov. as an appropriate generic name. (4). Rhizobium includes three species, Rhizobium meliloti,Rhizobium loti, and Rhizobium leguminosarum. Bradyrhizobium has only one species, Bradyrhizobium japonicum, and includes strains that are capable of nodulating lupines, soybeans, and certain other legumes (4).Recently, a new group of fast-growing soybean rhizobia was identified from soil and soybean nodules collected in the People's Republic of China by Chinese and American scientists (6, 25). The taxonomic status of these bacteria was uncertain, but they may represent a transitional group falling between the genera Rhizobium and Bradyrhizobium (4).Scholla and Elkan (13) proposed a new species name for this group, Rhizobium fredii, based mainly on deoxyribonucleic acid (DNA) hybridization comparisons of five strains of these bacteria with representatives of the genera Rhizobium and Brudyrhizobium and on a review of some of the characteristics of 11 strains of fast-growing soybean rhizobia.To extend the current information on the fast-growing soybean bacteria, we performed a numerical taxonomic study of a large number of strains. We propose that they be assembled in a new genus that includes two species. MATERIALS AND METHODSOrganisms. A total of 58 strains were used in this study (Table 1); 33 strains were fast-growing soybean rhizobia isolated from soil and soybean nodules collected in the People's Republic of China, and the other 25 strains were representatives of Rhizobium, ~r u d y r~i z o~i u m , and Agrobacterium.Features. Each of the 58 strains was characterized by determining 240 different coding features. The following features were considered: (i) utilization as sole carbon sources (0.1%) of cellobiose, D-mannose, salicin, ammonium tartrate, casein hydrolysate, fructose, sodium acetate, sorbose, riffinose, D-melezitose monohydrate, hexose 6-phos- (iv) tolerance to dyes (0.1%, wt/vol), such as sudan 1, erythrosin A, picrocarmine, safranine T, orcein, rosanilin, brilliant cresyl blue, gentian violet, bromthymol blue, bromophenol blue, methyl red, bromocresol purple, neutral red, auramine O.B.S, light gree...

A total of 63 strains of rhizobia isolated from Hainan Province, a tropical region of the People's Republic of China, and 27 representative strains belonging to the genera Rhizobium, Bradyrhizobium, and Agrobacterium were compared by performing numerical taxonomy, DNA hybridization, and DNA base composition analysis to determine the relationships among these rhizobia. The results indicated that the strains isolated from Hainan Province fell into two distinct phena, the slowly growing rhizobia and the fast-growing rhizobia. The slowly growing rhizobia, which formed three subphena that seemed to be three subspecies, are Bradyrhizobium japonicum strains. The fast-growing strains belong to the genus Rhizobium and might be further divided into three specific groups. Sometimes both slowly growing rhizobia and fast-growing rhizobia were isolated from host plants belonging to the same genus or species or even from the same nodule. There was no correlation between hosts and the distribution of rhizobia in the subphena. Isolates obtained from members of the same host genus or species fell into different groups or subgroups.The root nodule bacteria of tropical leguminous plants have been investigated by workers in many laboratories (1,(5)(6)(7)(8)10,23,25), but taxonomic studies of these bacteria have been sparse and have been limited to a few leguminous symbionts (16,19,24,27,28). Dreyfus and Dommergues (6) classified 13 Acacia species into three groups on the basis of effective nodulation with fast-growing and slowly growing tropical strains of rhizobia. The first group formed nodules effectively with slow growers; the second group formed effective nodules with fast growers; and the third group formed effective nodules with both fast and slow growers. Zhang et al. (27) obtained 12 clusters by using numerical taxonomy to study 97 strains isolated from root nodules of Acacia senagal, Prospopis chilensis, and other legumes. Thus, it is interesting to study the taxonomy of rhizobia isolated from a wide range of tropical legumes.In this work 63 rhizobial strains were isolated from various legumes, including trees, herbs, and vines, growing in different geographic regions of Hainan Province of the People's Republic of China, a tropical region south of latitude 20" N. These strains were compared with previously described species of rhizobia by performing numerical taxonomy, DNA composition, and DNA-DNA hybridization analysis. MATERIALS AND METHODSBacterial strains. A total of 90 strains were used in this study (Table 1); 27 of these strains were type and representative strains of Rhizobium, Bradyrhizobium, and Agrobacterium species, and 63 strains were isolated from root nodules collected in different geographic regions of Hainan Province. The purity of the cultures was assured by using single-colony isolates and checking colony morphology, bacterial morphology, and the Gram stain reaction. The identity of each strain was checked by performing a plant infection test with the original host plant.

Nine bacterial strains isolated from root nodules of Astragalus sinicus were compared with 41 reference strains, including the type strains of the type species of the genera Rhizobium, Bradyrhizobium, and Agrobacterium, by performing a numerical analysis of 200 phenotype features. Representative strains belonging to different clusters were further compared with similar bacteria by using data from gel electrophoresis of whole-cell proteins, DNA G + C content data, and DNA-DNA hybridization data. The rhizobial strains isolated from nodules of A . sinicus constitute a distinct homology group that is quite different from previously described Rhizobium, Bradyrhizobium, and Agrobacterium species and from strains isolated from other Astragalus species. We propose the name Rhizobium huakuii sp. nov. for the strains isolated from A . sinicus. Type strain CCBAU 2609 (= 103) has been deposited in the Culture Collection of Beijing Agricultural University, Beijing, People's Republic of China.Bacteria which form nitrogen-fixing nodules on leguminous plants are currently divided into the following four genera: Rhizobium, Bradyrhizobium, Azorhizobium, and Sinorhizobium (2, 5, 9). The genus Rhizobium comprises

Fifty-five Chinese isolates from nodules of Amorpha fruticosa were characterized and compared with the type strains of the species and genera of bacteria which form nitrogen-f ixing symbioses with leguminous host plants. A polyphasic approach, which included RFLP of PCR-amplif ied 165 rRNA genes, multilocus enzyme electrophoresis (MLEE), DNA-DNA hybridization, 165 rRNA gene sequencing, electrophoretic plasmid profiles, cross-nodulation and a phenotypic study, was used in the comparative analysis. The isolates originated from several different sites in China and they varied in their phenotypic and genetic characteristics. The majority of the isolates had moderate to slow growth rates, produced acid on YMA and harboured a 930 kb symbiotic plasmid (pSym). Five different RFLP patterns were identified among the 16s rRNA genes of all the isolates. Isolates grouped by PCR-RFLP of the 165 rRNA genes were also separated into groups by variation in MLEE profiles and by DNA-DNA hybridization. A representative isolate from each of these DNA homology groups had a separate position in a phylogenetic tree as determined from sequencing analysis of the 165 rRNA genes. A new species, Mesorhizobium amorphae, is proposed for the majority of the isolates, which belonged to a moderately slow-to slow-growing, acid-producing group based upon their distinct phylogenetic position, their unique electrophoretic type, their low DNA homology with reference strains representing the species within the genus Mesorhizobium and their distinct phenotypic features. Strain ACCC 19665 was chosen as the type strain for M. amorphae sp. nov.