Rhizobium pisi sv. trifolii K3.22 harboring nod genes of the Rhizobium leguminosarum sv. trifolii cluster

The taxonomic status of the Rhizobium sp. K3.22 clover nodule isolate was studied by multilocus sequence analysis (MLSA) of 16S rRNA and six housekeeping chromosomal genes, as well as by a subsequent phylogenic analysis. The results revealed full congruence with the Rhizobium pisi DSM 30132^T core genes, thus supporting the same taxonomic position for both strains. However, the K3.22 plasmid symbiosis nod genes demonstrated high sequence similarity to Rhizobium leguminosarum sv. trifolii, whereas the R. pisi DSM 30132^Tnod genes were most similar to R. leguminosarum sv. viciae. The strains differed in the host range nodulation specificity, since strain K3.22 effectively nodulated red and white clover but not vetch, in contrast to R. pisi DSM 30132^T, which effectively nodulated vetch but was not able to nodulate clover. Both strains had the ability to form nodules on pea and bean but they differed in bean cultivar specificity. The R. pisi K3.22 and DSM 30132^T strains might provide evidence for the transfer of R. leguminosarum sv. trifolii and sv. viciae symbiotic plasmids occurring in natural soil populations.     

Genomic insight into the taxonomy of Rhizobium genospecies that nodulate Phaseolus vulgaris

Due to the wide cultivation of bean (Phaseolus vulgaris L.), rhizobia associated with this plant have been isolated from many different geographical regions. In order to investigate the species diversity of bean rhizobia, comparative genome sequence analysis was performed in the present study for 69 Rhizobium strains mainly isolated from root nodules of bean and clover (Trifolium spp.). Based on genome average nucleotide identity, digital DNA:DNA hybridization, and phylogenetic analysis of 1,458 single-copy core genes, these strains were classified into 28 clusters, consistent with their species definition based on multilocus sequence analysis (MLSA) of atpD, glnII, and recA. The bean rhizobia were found in 16 defined species and nine putative novel species; in addition, 35 strains previously described as Rhizobium etli, Rhizobium phaseoli, Rhizobium vallis, Rhizobium gallicum, Rhizobium leguminosarum and Rhizobium spp. should be renamed. The phylogenetic patterns of symbiotic genes nodC and nifH were highly host-specific and inconsistent with the genomic phylogeny. Multiple symbiovars (sv.) within the Rhizobium species were found as a common feature: sv. phaseoli, sv. trifolii and sv. viciae in Rhizobium anhuiense; sv. phaseoli and sv. mimosae in Rhizobium sophoriradicis/R. etli/Rhizobium sp. III; sv. phaseoli and sv. trifolii in Rhizobium hidalgonense/Rhizobium acidisoli; sv. phaseoli and sv. viciae in R. leguminosarum/Rhizobium sp. IX; sv. trifolii and sv. viciae in Rhizobium laguerreae. Thus, genomic comparison revealed great species diversity in bean rhizobia, corrected the species definition of some previously misnamed strains, and demonstrated the MLSA a valuable and simple method for defining Rhizobium species.     

The analysis of core and symbiotic genes of rhizobia nodulating Vicia from different continents reveals their common phylogenetic origin and suggests the distribution of Rhizobium leguminosarum strains together with Vicia seeds

In this work, we analysed the core and symbiotic genes of rhizobial strains isolated from Vicia sativa in three soils from the Northwest of Spain, and compared them with other Vicia endosymbionts isolated in other geographical locations. The analysis of rrs, recA and atpD genes and 16S–23S rRNA intergenic spacer showed that the Spanish strains nodulating V. sativa are phylogenetically close to those isolated from V. sativa and V. faba in different European, American and Asian countries forming a group related to Rhizobium leguminosarum. The analysis of the nodC gene of strains nodulating V. sativa and V. faba in different continents showed they belong to a phylogenetically compact group indicating that these legumes are restrictive hosts. The results of the nodC gene analysis allow the delineation of the biovar viciae showing a common phylogenetic origin of V. sativa and V. faba endosymbionts in several continents. Since these two legume species are indigenous from Europe, our results suggest a world distribution of strains from R. leguminosarum together with the V. sativa and V. faba seeds and a close coevolution among chromosome, symbiotic genes and legume host in this Rhizobium–Vicia symbiosis.     

Rhizobium sophorae is the dominant rhizobial symbiont of Vicia faba L. In North China

Faba bean (Vicia faba L.) is a major introduced grain-legume crop cultivated in China. In this study, rhizobia that nodulated faba bean grown in soils from three sites in North China (Hebei Province) were isolated and characterized. Firstly, isolates were categorized into genotypes by ribosomal IGS PCR-RFLP analysis, then representatives of the different IGS genotypes were further identified by phylogenetic analyses of 16S rRNA, housekeeping (atpD, recA) and nodulation (nodC) gene sequences. Rhizobial distribution based on the IGS genotype was related to the different soil physicochemical features by redundancy analysis. IGS typing and phylogenetic analyses of 16S rRNA and concatenated housekeeping gene sequences affiliated the 103 rhizobial strains isolated into four Rhizobium species/genospecies. A total of 69 strains of 3 IGS types were assigned to R. sophorae, 20 isolates of 5 IGS types to R. changzhiense and 9 isolates of 3 IGS types to R. indicum. The representative strain of the five remaining isolates (1 IGS type) was clearly separated from all Rhizobium type strains and was most closely related to defined genospecies according to the recently described R. leguminosarum species complex. Rhizobium sophorae strains (67% of total isolates) were common in all sites and shared an identical nodC sequence typical of faba bean symbionts belonging to symbiovar viciae. In this first study of rhizobia nodulating faba bean in Hebei Province, China, R. sophorae was found to be the dominant symbiont in contrast to other countries.     

Microsymbionts of Phaseolus vulgaris in acid and alkaline soils of Mexico

In order to investigate bean-nodulating rhizobia in different types of soil, 41 nodule isolates from acid and alkaline soils in Mexico were characterized. Based upon the phylogenetic studies of 16S rRNA, atpD, glnII, recA, rpoB, gyrB, nifH and nodC genes, the isolates originating from acid soils were identified as the phaseoli symbiovar of the Rhizobium leguminosarum-like group and Rhizobium grahamii, whereas the isolates from alkaline soils were defined as Ensifer americanum sv. mediterranense and Rhizobium radiobacter. The isolates of “R. leguminosarum” and E. americanum harbored nodC and nifH genes, but the symbiotic genes were not detected in the four isolates of the other two species. It was the first time that “R. leguminosarum” and E. americanum have been reported as bean-nodulating bacteria in Mexico. The high similarity of symbiotic genes in the Rhizobium and Ensifer populations showed that these genes had the same origin and have diversified recently in different rhizobial species. Phenotypic characterization revealed that the “R. leguminosarum” population was more adapted to the acid and low salinity conditions, while the E. americanum population preferred alkaline conditions. The findings of this study have improved the knowledge of the diversity, geographic distribution and evolution of bean-nodulating rhizobia in Mexico.     

Phylogenetic diversity based on <Emphasis Type="Italic">rrs,atpD, recA</Emphasis> genes and 16S–23S intergenic sequence analyses of rhizobial strains isolated from <Emphasis Type="Italic">Vicia faba</Emphasis> and <Emphasis Type="Italic">Pisum sativum</Emphasis> in Peru

In this study 17 isolates from effective nodules of Vicia faba and Pisum sativum var. macrocarpum growing in different soils from Peru were isolated and characterized. The isolates, presenting 11 different RAPD profiles, were distributed in three groups on the basis of their 16S-RFLP patterns. The 16S rRNA gene sequences of strains from 16S-RFLP groups I, II and III were closely related (identities higher than 99.5%) to Rhizobium leguminosarum bv. trifolii DSM 30141 (=ATCC 14480), R. leguminosarum bv. viciae DSM 30132^T and Rhizobium etli CFN42^T (=USDA 9032^T), respectively. The analysis of the 16S–23S intergenic spacer (ITS) and two housekeeping genes, atpD and recA, confirmed the identification of strains from group I, however those from groups II and III were phylogenetically divergent to strains DSM 30132^T and CFN42^T. These results support the fact that the 16S rRNA gene is not adequate for identification at species level within genus Rhizobium and suggest the existence of putative new species within the phylogenetic group of R. leguminosarum. They also confirm the need of a taxonomic revision of R. leguminosarum since the reference strains of the three biovars included in this study are phylogenetically divergent according to their ITS, atpD and recA gene sequences.     

Characterization of Pisum sativum and Vicia faba microsymbionts in Morocco and definition of symbiovar viciae in Rhizobium acidisoli

In this work, we analyzed the diversity of seventy-six bacteria isolated from Pea and faba bean nodules in two regions of Morocco. The molecular diversity was realized using the analysis of the sequences of 16S rRNA and six housekeeping genes (recA, glnII, atpD, dnaK, rpoB and gyrB) and two symbiotic genes (nodA and nodC).The phylogeny of the 16S rRNA gene sequences revealed that all strains belong to the genus Rhizobium, being related to the type strains of R. leguminosarum, R. laguerreae, R. indigoferae, R. anhuiense and R. acidisoli. The housekeeping genes phylogenies showed that some strains formed a subclade distinct from the rhizobial species that usually nodulate Vicia faba and Pisum sativum which are closely related to R. acidisoli FH23 with sequence similarity of 98.3%.Analysis of the PGPR activities of the different isolates showed that the strains related to R. laguerreae were able to solubilize phosphates and to produce siderophores and auxin phytohormone. However, R. acidisoli strain F40D2 was unable to solubilize phosphates although they produce siderophores and IAA.The phylogenetic analysis of the nodA and nodC sequences showed that all isolated strains were closely related with the strains of symbiovar viciae. The nodulation tests confirmed the ability to nodulate V. faba and P. sativum but not Cicer arietinum or Phaseolus vulgaris. Hence, in Morocco P. sativum is nodulated by R. laguerreae; whereas V. faba is nodulated by R. laguerreae and the symbiovar viciae of R. acidisoli which has been not previously described in this species.     

Phylogenetic multilocus sequence analysis of native rhizobia nodulating faba bean (Vicia faba L.) in Egypt

The taxonomic diversity of forty-two Rhizobium strains, isolated from nodules of faba bean grown in Egypt, was studied using 16S rRNA sequencing, multilocus sequence analyses (MLSA) of three chromosomal housekeeping loci and one nodulation gene (nodA). Based on the 16S rRNA gene sequences, most of the strains were related to Rhizobium leguminosarum, Rhizobium etli, and Rhizobium radiobacter (syn. Agrobacterium tumefaciens). A maximum likelihood (ML) tree built from the concatenated sequences of housekeeping proteins encoded by glnA, gyrB and recA, revealed the existence of three distinct genospecies (I, II and III) affiliated to the defined species within the genus Rhizobium/Agrobacterium. Seventeen strains in genospecies I could be classified as R. leguminosarum sv. viciae. Whereas, a single strain of genospecies II was linked to R. etli. Interestingly, twenty-four strains of genospecies III were identified as A. tumefaciens. Strains of R. etli and A. tumefaciens have been shown to harbor the nodA gene and formed effective symbioses with faba bean plants in Leonard jar assemblies. In the nodA tree, strains belonging to the putative genospecies were closely related to each other and were clustered tightly to R. leguminosarum sv. viciae, supporting the hypothesis that symbiotic and core genome of the species have different evolutionary histories and indicative of horizontal gene transfer among these rhizobia.     

Phylogenetic diversity of Bradyrhizobium strains nodulating Calicotome spinosa in the Northeast of Algeria

Fifty-two slow-growing strains were isolated from root nodules of Calicotome spinosa grown in the Northeast of Algeria and grouped in 24 rep-PCR clusters. One representative strain for each profile was further phylogenetically characterized. The nearly complete 16S rRNA gene sequence indicated that all strains were affiliated to Bradyrhizobium. Multi-Locus Sequence Analysis (MLSA) of the atpD, glnII and recA genes and of the 16S-23S rRNA internal transcribed spacer (ITS) showed that these strains formed four divergent clusters: one close to Bradyrhizobium canariense and Bradyrhizobium lupini and three others separate from all the described species, representing three putative new Bradyrhizobium species. A phylogenetic analysis based on the nodC gene sequence affiliated the strains to either of the two symbiovars, genistearum or retamae.     

A genetic discontinuity in root-nodulating bacteria of cultivated pea in the Indian trans-Himalayas

Evolutionary relationships of 120 root‐nodulating bacteria isolated from the nodules of Pisum sativum cultivated at 22 different locations of the trans‐Himalayan valleys of Lahaul and Spiti in the state of Himachal Pradesh of India were studied using 16S rRNA gene PCR‐RFLP, ERIC‐PCR, sequencing of 16S rRNA, atpD, recA, nodC and nifH genes, carbon‐source utilization pattern (BIOLOG?), and whole‐cell fatty acid profiling. The results demonstrated that all isolates belonged to Rhizobium leguminosarum symbiovar viciae (Rlv). Isolates from the two valleys were clearly separated on the basis of ERIC fingerprints, carbon‐source utilization pattern, and whole‐cell fatty acid methyl esters. Phylogenetic analysis of atpD, recA, nodC and nifH genes revealed a common Rlv sublineage in Spiti valley. Lahaul valley isolates were represented by three sequence types of atpD and recA genes, and four sequence types of nodC and nifH genes. Genotypes from the two valleys were completely distinct, except for two Lahaul isolates that shared nodC and nifH sequences with Spiti isolates but were otherwise more similar to other Lahaul isolates. Isolates from the two highest Spiti valley sites (above 4000 m) had a distinctive whole‐cell fatty acid profile. Spiti valley isolates are closely related to Rlv sublineages from Xinjiang and Shanxi provinces in China, while Lahaul valley isolates resemble cosmopolitan strains of the western world. The high mountain pass between these valleys represents a boundary between two distinct microbial populations.     

Near-full length sequencing of 16S rDNA and RFLP indicates that Rhizobium etli is the dominant species nodulating Egyptian winter Berseem clover (Trifolium alexandrinum L.)

Egyptian winter Berseem clover (EWBC) is one of the main important forage legume crops in Egypt that is used for animal feeding in winter and it occupies about 2.5 million feddans (Feddan = 4200 m²) in winter agricultural rotation systems. Forty-eight rhizobial isolates that nodulated this legume host from different geographical regions within Egypt were isolated. RFLP analyses of 16S rDNA (1.5 kb) and whole ribosomal DNA (5 kb), the sequencing of 16S rDNA, and the sequencing of nodC, nifH and house keeping genes were used to identify these isolates. The RFLP analysis of 16S rDNA (1.5 kb) among 15 representative strains with three enzymes generated two genotypes. The largest genotype was similar to Rhizobium etli CFN42T (93.33%) except for strain 902 that failed to re-nodulate EWBC. RFLP analysis of complete ribosomal DNA (5 kb) produced five genotypes. The majority of tested strains shared the genotype with R. etli CFN42T (53.33%). Only one strain (1002) shared the genotype with Rhizobium leguminosarum sv. trifolii 3023. The other four strains were comprised of two unique genotypes. Phylogenetic analysis of 16S rDNA sequences revealed that seven representative strains could be divided into two genetic clusters sharing the ancestral clad with R. etli CFN42T. A phylogenetic tree based on nodC gene sequence confirmed that all the examined strains shared the genetic lineage with R. leguminosarum sv. trifolii WSM1325. The phylogenetic trees of house keeping genes are supported strongly the identification of majority of strains as a novel symbiovar of R. etli with new lineages.     

Rhizobium laguerreae is the main nitrogen-fixing symbiont of cultivated lentil (Lens culinaris) in Morocco

Genetic diversity and population structure of 268 Lens culinaris symbiotic rhizobia collected from 40 cultivated fields in the main lentil production regions in Morocco were estimated. Three chromosomal housekeeping genes (recA, glnII and atpD) and one common symbiotic gene (nodC) were sequenced and analyzed in order to identify the local symbionts of lentil. The molecular phylogeny of the concatenated housekeeping genes clustered more than 95% of the isolates in one main clade together with Rhizobium laguerreae species. R. laguerreae represents the main symbiont of cultivated lentil in Morocco and, for the first time, a large sample of individuals is obtained for this species. There is a significant and high genetic differentiation of bacterial populations among the four regions for their symbiotic gene, and much lower for their housekeeping genes. The reasons why R. laguerreae is so frequently recovered in our study is discussed.     

分离自印度洋深海沉积物的部分链霉菌分离株多位点序列分析

【目的】采用多位点序列分析方法,研究印度洋3 000 m以下深海沉积物中分离得到的16S rRNA基因比对高度相似的链霉菌菌株的种间系统发育关系,同时探讨各管家基因及多基因聚类分析后的种间区分能力。【方法】以分离自印度洋深海沉积物的7株Streptomyces albidoflavus,11株Streptomyces cavourensis,16株Streptomyces pratensis为研究对象,以16S rRNA、atpD、recA和rpoB基因片段为标记,通过PCR扩增、测序,获得序列。同时从NCBI上下载5株S.pratensis上述4个基因的序列,将所有序列在MLST网站进行比对,并构建系统进化树进行比较。【结果】S.pratensis各菌株种内比较发现,16S rRNA基因构建的系统进化树中相同基因型的菌株没有聚在一起,系统进化树不稳定,区分度不高。其余3个构建的系统进化树稳定,菌株的聚类关系与MLST数据库得到的基因型一致。同时,多基因聚类分析后将菌株分为6个类群。在3个种的种间多位点序列比较中,除区分度明显增加、进化树更加稳定以外,还发现rec A基因进化上比较特殊的菌株。【结论】多位点序列分析将实验菌株分为很多不同的类型,成功地将所分离的链霉菌进行了更细的分类,同时也找到部分菌株在个别基因上差异较大。此方法可以用于相近种的快速鉴定。     

Phaseolus vulgaris seed-borne endophytic community with novel bacterial species such as Rhizobium endophyticum sp. nov.

The bacterial endophytic community present in different Phaseolus vulgaris (bean) cultivars was analyzed by 16S ribosomal RNA gene sequences of cultured isolates derived from surface disinfected roots and immature seeds. Isolated endophytes from tissue-macerates belonged to over 50 species in 24 different genera and some isolates from Acinetobacter, Bacillus, Enterococcus, Nocardioides, Paracoccus, Phyllobacterium, and Sphingomonas seem to correspond to new lineages. Phytate solubilizing bacteria were identified among Acinetobacter, Bacillus and Streptomyces bean isolates, phytate is the most abundant reserve of phosphorus in bean and in other seeds. Endophytic rhizobia were not capable of forming nodules. A novel rhizobial species Rhizobium endophyticum was recognized on the basis of DNA–DNA hybridization, sequence of 16S rRNA, recA, rpoB, atpD, dnaK genes, plasmid profiles, and phenotypic characteristics. R. endophyticum is capable of solubilizing phytate, the type strain is CCGE2052 (ATCC BAA-2116; HAMBI 3153) that became fully symbiotic by acquiring the R. tropici CFN299 symbiotic plasmid.     

Genetic diversity of Vicia faba L. and Pisum sativum L. nodulating rhizobia in the central Black Sea region of Turkey

In this study, we obtained a total of 60 rhizobial isolates from root nodules of Vicia faba L. (n = 30) and Pisum sativum L. (n = 30) grown in the Central Black Sea region of Turkey. The 16S rDNA PCR-RFLP analysis with enzymes CfoI, HinfI, NdeII and MspI revealed a single pattern. Moreover, nucleotide sequence phylogenies based on both the 16S rDNA and recA suggested that these isolates belonged to Rhizobium leguminosarum. Phylogenetic analysis showed that some of our V. faba L.-originated isolates were closely related, indicating molecular evidence for the selection of some special R. leguminosarum bv. viciae isolates by V. faba L., as suggested in previous studies. Network analysis based on recA sequences revealed a common evolutionary history for Turkish, European, North and South American, and Jordanian R. leguminosarum bv. viciae isolates. We isolated four haplotypes using nodA and nifH nucleotide sequence data, i.e. four types of sym plasmids. Two of these types were common to rhizobial isolates from both V. faba L. and P. sativum L., indicating that nodulation factors may not be the mechanism for selection of the special R. leguminosarum bv. viciae populations by V. faba L.     

Rhizobium indicum sp. nov., isolated from root nodules of pea (Pisum sativum) cultivated in the Indian trans-Himalayas

Three strains of rhizobia isolated from effective root nodules of pea (Pisum sativum L.) collected from the Indian trans-Himalayas were characterized using 16S rRNA, atpD and recA genes. Phylogeny of the 16S rRNA genes revealed that the newly isolated strains were members of the genus Rhizobium with ≥99.9% sequence similarity to the members within the “Rhizobium leguminosarum” group. Phylogenetic analyses based on the concatenated sequences of atpD and recA gene, and 92 core genes extracted from the genome sequences indicated that strains JKLM 12A2^T and JKLM 13E are grouped as a separate clade closely related to R. laguerreae FB206^T. In contrast, the strain JKLM 19E was placed with “R. hidalgonense” FH14^T. Whole-genome average nucleotide identity (ANI) values were 97.6% within strains JKLM 12A2^T and JKLM 13E, and less than 94% with closely related species. The digital DNA-DNA hybridization (dDDH) values were 81.45 within the two strains and less than 54.8% to closely related species. The major cellular fatty acids were C_18:1w7c in summed feature 8, C_{14:0 3OH}/C_{16:1 iso I} in summed feature 2, and C_18:0. The DNA G + C content of JKLM 12A2^T and JKLM 13E was 60.8 mol%. The data on genomic, chemotaxonomic, and phenotypic characteristics indicates that the strains JKLM 12A2^T and JKLM 13E represent a novel species, Rhizobium indicum sp. nov. The type strain is JKLM 12A2^T (= MCC 3961^T = KACC 21380^T = JCM 33658^T). However, the strain JKLM 19E represents a member of “R. hidalgonense” and the symbiovar viciae.     

Phenotypic and genetic diversity of Moroccan rhizobia isolated from Vicia faba and study of genes that are likely to be involved in their osmotolerance

Rhizobia are symbiotic nitrogen-fixing bacteria in root nodules of legumes. In Morocco, faba bean (Vicia faba L.), which is the main legume crop cultivated in the country, is often grown in marginal soils of arid and semi-arid regions. This study examines the phenotypic diversity of rhizobia nodulating V. faba isolated from different regions in Morocco for tolerance to some abiotic stresses. A total of 106 rhizobia strains isolated from nodules were identified at the species level by analysing 16S rDNA. Additionally, for selected strains recA, otsA, kup and nodA fragments were sequenced. 102 isolates are likely to belong to Rhizobium leguminosarum or R. laguerreae and 4 isolates to Ensifer meliloti. All strains tolerating salt concentrations of 428 or 342 mM NaCl as well as 127 or 99 mM Na2SO4 were highly resistant to alkaline conditions (pH 10) and high temperature (44 °C). Three strains: RhOF4 and RhOF53 (both are salt-tolerant) and RhOF6 (salt-sensitive) were selected to compare the influence of different levels of salt stress induced by NaCl on growth and on trehalose and potassium accumulation. We find a direct correlation between the trehalose contents of the rhizobial strains and their osmotolerance.     

Analysis of rhizobial endosymbionts of Vicia,Lathyrus and Trifolium species used to maintain mountain firewalls in Sierra Nevada National Park (South Spain)

Forest fires lead to the annual disappearance of many natural formations that require the creation of firewall areas. They can be maintained by enriching their pastures with attractive plants for grazing livestock, mainly legumes, which have a high protein content and low dependence on N fertilizers due to their ability to establish nitrogen-fixing symbiosis with rhizobia. In this study, the rhizobia isolated from the nodules of six legumes from the genera Vicia, Lathyrus and Trifolium were analysed in a firewall zone established in Lanjarón (Granada) close to the Sierra Nevada National Park (Spain). The results showed a high genetic diversity of the isolated strains that had 3, 16, 14 and 13 different types of rrs, recA, atpD and glnII genes, respectively. All strains were phylogenetically close to the species from the Rhizobium leguminosarum group, although they were not identified as any of them. The isolated strains belonged to the symbiovars viciae and trifolii but high phylogenetic diversity was found within both symbiovars, since there were 16 and 14 nodC gene types, respectively. Some of these strains clustered with strains isolated in other countries and continents, but others formed atpD, recA, glnII and nodC clusters and lineages only found to date in this study.     

Distinctive Mesorhizobium populations associated with Cicer arietinum L. in alkaline soils of Xinjiang, China

Background and aims

Rhizobia associated with chickpea in the main chickpea production zone of Xinjiang, China have never been investigated. Here, we present the first systematic investigation of these rhizobia’s genetic diversity and symbiotic interactions with their host plant.

Methods

Ninety-five isolates obtained from chickpea nodules in eight alkaline-saline (pH?8.24–8.45) sites in Xinjiang were characterized by nodulation test, symbiotic gene analysis, PCR-based restriction fragment length polymorphism (RFLP) of the 16S rRNA gene and 16S–23S rRNA intergenic spacer (IGS), BOX-PCR, phylogenies of 16S rRNA and housekeeping genes (atpD, recA and glnII), multilocus sequence analysis (MLSA) and DNA–DNA hybridization.

Results

All 95 isolates were identified within the genus of Mesorhizobium. Similarities less than 96.5% in MLSA and DNA–DNA hybridization values (<50%) between the new isolates and the defined Mesorhizobium species, and high similarities (>98%) of symbiotic genes (nodC and nifH) with those of the well studied chickpea microsymbioints Mesorhizobium ciceri and Mesorhizobium mediterraneum were found.

Conclusions

Chickpea rhizobia in alkaline-saline soils of Xinjiang, China, form a population distinct from the defined Mesorhizobium species. All these chickpea rhizobia in Xinjiang harbored symbiotic genes highly similar to the type strains of two well-studied chickpea rhizobia, M. ciceri and M. mediterraneum, evidencing the possible lateral transfer of symbiotic genes among these different rhizobial species. On the other hand, chickpea may strongly select rhizobia with a unique symbiotic gene background.     

Classification of Streptomyces phylogroup pratensis (Doroghazi and Buckley, 2010) based on genetic and phenotypic evidence,and proposal of Streptomyces pratensis sp. nov.

The Streptomyces phylogroup pratensis (Doroghazi and Buckley, 2010) contains isolates obtained from grassy fields, as well as Streptomyces flavogriseus ATCC 33331 and strain CGMCC 4.1868. This latter strain was received as Streptomyces griseoplanus but was subsequently found to be mislabeled, and S. flavogriseus ATCC 33331 (=IAF-45-CD) was shown to be clearly distinct from the type strain S. flavogriseus ATCC 25452^T (=CGMCC 4.1884^T). In order to evaluate the taxonomic position of phylogroup pratensis further, sequences of the 16S rRNA gene and five protein-coding housekeeping genes (atpD, gyrB, recA, rpoB and trpB) were determined for six strains of the phylogroup and type strains of 19 related species, which were selected by a BLAST search based on the sequences of the phylogroup. The 16S rRNA gene sequences for the phylogroup were identical to those of eight species belonging to cluster I of the S. griseus clade. However, in all the individual protein-coding gene and MLSA phylogenies, the phylogroup strains without exception formed an obviously distinct cluster that could be equated with a new species status. The phylogenetic evidence for the new species assignment was also supported by corresponding DNA–DNA hybridization values and by phenotypic characteristics. It is therefore proposed that the phylogroup should be classified as Streptomyces pratensis sp. nov., and the type strain is ch24^T (=CGMCC 4.6829^T = NRRL B-24916^T).