Skip to main content
SpringerLink
Log in

Bradyrhizobium sacchari sp. nov., a legume nodulating bacterium isolated from sugarcane roots

  • Original Paper
  • Published:
Archives of Microbiology Aims and scope Submit manuscript

Abstract

Members of the genus Bradyrhizobium are well-known as nitrogen-fixing microsymbionts of a wide variety of leguminous species, but they have also been found in different environments, notably as endophytes in non-legumes such as sugarcane. This study presents a detailed polyphasic characterization of four Bradyrhizobium strains (type strain BR 10280T), previously isolated from roots of sugarcane in Brazil. 16S rRNA sequence analysis, multilocus sequence analysis (MLSA) and analysis of the 16S-23S rRNA internal transcribed spacer showed that these strains form a novel clade close to, but different from B. huanghuaihaiense strain CCBAU 23303T. Average nucleotide identity (ANI) analyses confirmed that BR 10280T represents a novel species. Phylogenetic analysis based on nodC gene sequences also placed the strains close to CCBAU 23303T, but different from this latter strain, the sugarcane strains did not nodulate soybean, although they effectively nodulated Vigna unguiculata, Cajanus cajan and Macroptilium atropurpureum. Physiological traits are in agreement with the placement of the strains in the genus Bradyrhizobium as a novel species for which the name Bradyrhizobium sacchari sp. nov. is proposed.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • Baldani JI, Baldani VL (2005) History on the biological nitrogen fixation research in graminaceous plants: special emphasis on the Brazilian experience. An Acad Bras Cienc 77:549–579

    Article  CAS  PubMed  Google Scholar 

  • Baldani JI, Reis VM, Videira SS, Boddey LH, Baldani VLD (2014) The art of isolating nitrogen-fixing bacteria from non-leguminous plants using N-free semi-solid media: a practical guide for microbiologists. Plant Soil 384:413–431

    Article  CAS  Google Scholar 

  • Cardinale M, Brusetti L, Quatrini P, Borin S, Puglia AM, Rizzi A, Zanardini E, Sorlini C, Corselli C, Daffonchio D (2004) Comparison of different primer sets for use in automated ribosomal intergenic spacer analysis of complex bacterial communities. Appl Environ Microbiol 70:6147–6156

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chaintreuil C, Giraud E, Prin Y, Lorquin J, Bâ A, Gillis M, Lajudie P, Dreyfus B (2000) Photosynthetic bradyrhizobia are natural endophytes of the African wild rice Oryza breviligulata. Appl Environ Microbiol 66:5437–5447

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Delamuta JRM, Ribeiro RA, Ormeño-Orrillo E, Melo IS, Martínez-Romero E, Hungria M (2013) Polyphasic evidence supporting the reclassification of Bradyrhizobium japonicum group Ia strains as Bradyrhizobium diazoefficiens sp. nov. Int J Syst Evol Microbiol 63:3342–3351

    Article  CAS  PubMed  Google Scholar 

  • Delamuta JRM, Ribeiro RA, Ormeño-Orrillo E, Parma MM, Melo IS, Martínez-Romero E, Hungria M (2015) Bradyrhizobium tropiciagri sp. nov. and Bradyrhizobium embrapense sp. nov., nitrogen-fixing symbionts of tropical forage legumes. Int J Syst Evol Microbiol 65:4424–4433

    Article  CAS  PubMed  Google Scholar 

  • Delamuta JRM, Ribeiro RA, Araújo JLS, Rouws LFM, Zilli JE, Parma MM, Melo IS, Hungria M (2016) Bradyrhizobium stylosanthis sp. nov., comprising nitrogen-fixing symbionts isolated from nodules of the tropical forage legume Stylosanthes spp. Int J Syst Evol Microbiol. doi:10.1099/ijsem.0.001148

    PubMed  Google Scholar 

  • Durán D, Rey L, Navarro A, Busquets A, Imperial J, Ruiz-Argüeso T (2014) Bradyrhizobium valentinum sp. nov., isolated from effective nodules of Lupinus mariae-josephae, a lupine endemic of basic-lime soils in Eastern Spain. Syst Appl Microbiol 37:336–341

    Article  PubMed  Google Scholar 

  • Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32:1792–1797

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Felsenstein J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–791

    Article  PubMed  Google Scholar 

  • Fischer D, Pfitzner B, Schmid M, Simões-Araújo JL, Reis VM, Pereira W, Ormeño-Orillo E, Hai B, Hofmann A, Martinez-Romero E (2012) Molecular characterisation of the diazotrophic bacterial community in uninoculated and inoculated field-grown sugarcane (Saccharum sp.). Plant Soil 356:83–99

    Article  CAS  Google Scholar 

  • Galtier N, Gouy M, Gautier C (1996) SEAVIEW and PHYLO_WIN: two graphic tools for sequence alignment and molecular phylogeny. Comp Appl Biosci 12:543–548

    CAS  PubMed  Google Scholar 

  • Goris J, Konstantinidis KT, Klappenbach JA, Coenye T, Vandamme P, Tiedje JM (2007) DNA–DNA hybridization values and their relationship to whole-genome sequence similarities. Int J Syst Evol Microbiol 57:81–91

    Article  CAS  PubMed  Google Scholar 

  • Gutierrez-Zamora ML, Martınez-Romero E (2001) Natural endophytic association between Rhizobium etli and maize (Zea mays L.). J Biotechnol 91:117–126

    Article  CAS  PubMed  Google Scholar 

  • Hallmann J, Quadt-Hallmann A, Mahaffee WF, Kloepper JW (1997) Bacterial endophytes in agricultural crops. Can J Microbiol 43:895–914

    Article  CAS  Google Scholar 

  • Konstantinidis KT, Tiedje JM (2005) Genomic insights that advance the species definition for prokaryotes. Proc Natl Acad Sci USA 102:2567–2572

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lee I, Kim YO, Park SC, Chun J (2016) OrthoANI: an improved algorithm and software for calculating average nucleotide identity. Int J Syst Evol Microbiol 66:1100–1103

    Article  CAS  Google Scholar 

  • Lima E, Boddey RM, Döbereiner J (1987) Quantification of biological nitrogen fixation associated with sugar cane using a 15N aided nitrogen balance. Soil Biol Biochem 19:165–170

    Article  CAS  Google Scholar 

  • Martens M, Delaere M, Coopman R, De Vos P, Gillis M, Willems A (2007) Multilocus sequence analysis of Ensifer and related taxa. Int J Syst Evol Microbiol 57:489–503

    Article  CAS  PubMed  Google Scholar 

  • Menna P, Barcellos FG, Hungria M (2009) Phylogeny and taxonomy of a diverse collection of Bradyrhizobium strains based on multilocus sequence analysis of the 16S rRNA gene, ITS region and glnII, recA, atpD and dnaK genes. Int J Syst Evol Microbiol 59:2934–2950

    Article  CAS  PubMed  Google Scholar 

  • Norris DO, Date RA (1976) Legume bacteriology. In: Shaw NH, Bryan WW (eds) Tropical pasture research; principles and methods. Commonwealth Agricultural Bureaux, Slough, UK, pp 134–174

    Google Scholar 

  • Radl V, Simões-Araújo JL, Leite J, Passos SR, Martins LMV, Xavier GR, Rumjanek NG, Baldani JI, Zilli JE (2014) Microvirga vignae sp. nov., a root nodule symbiotic bacterium isolated from cowpea grown in semi-arid Brazil. Int J Syst Evol Microbiol 64:725–730

    Article  PubMed  Google Scholar 

  • Richter M, Rosselló-Móra R (2009) Shifting the genomic gold standard for the prokaryotic species definition. Proc Natl Acad Sci USA 106:19126–19131

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rivas R, Martens M, De Lajudie P, Willems A (2009) Multilocus sequence analysis of the genus Bradyrhizobium. Syst Appl Microbiol 32:101–110

    Article  CAS  PubMed  Google Scholar 

  • Rosenblueth M, Martínez-Romero E (2004) Rhizobium etli maize populations and their competitiveness for root colonization. Arch Microbiol 181:337–344

    Article  CAS  PubMed  Google Scholar 

  • Rouws LFM, Leite J, Matos GF, Zilli JE, Coelho MRR, Xavier GR, Fisher D, Hartmann A, Reis VM, Baldani JI (2014) Endophytic Bradyrhizobium spp. isolates from sugarcane obtained through different culture strategies. Env Microbiol Rep 6:354–363

    Article  CAS  Google Scholar 

  • Ruschel AP, Henis Y, Salati E (1975) Nitrogen-15 tracing of N-fixation with soil-grown sugarcane seedlings. Soil Biol Biochem 7:181–182

    Article  CAS  Google Scholar 

  • Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425

    CAS  PubMed  Google Scholar 

  • Sarita S, Sharma PK, Priefer UB, Prell J (2005) Direct amplification of rhizobial nodC sequences from soil total DNA and comparison to nodC diversity of root nodule isolates. FEMS Microbiol Ecol 54:1–11

    Article  CAS  PubMed  Google Scholar 

  • Stępkowski T, Moulin L, Krzyżańska A, McInnes A, Law IJ, Howieson J (2005) European origin of Bradyrhizobium populations infecting lupins and serradella in soils of Western Australia and South Africa. Appl Environ Microbiol 71:7041–7052

    Article  PubMed  PubMed Central  Google Scholar 

  • Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol 30:2725–2729

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Thaweenut N, Hachisuka Y, Ando S, Yanagisawa S, Yoneyama T (2011) Two seasons’ study on nifH gene expression and nitrogen fixation by diazotrophic endophytes in sugarcane (Saccharum spp. hybrids): expression of nifH genes similar to those of rhizobia. Plant Soil 338:435–449

    Article  CAS  Google Scholar 

  • Tighe SW, De Lajudie P, Dipietro K, Lindström K, Nick G, Jarvis BD (2000) Analysis of cellular fatty acids and phenotypic relationships of Agrobacterium, Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium species using the Sherlock Microbial Identification System. Int J Syst Evol Microbiol 50:787–801

    Article  CAS  PubMed  Google Scholar 

  • Urquiaga S, Cruz KH, Boddey RM (1992) Contribution of nitrogen fixation to sugar cane: nitrogen-15 and nitrogen-balance estimates. Soil Sci Soc America J 56:105–114

    Article  Google Scholar 

  • Urquiaga S, Xavier RP, de Morais RF, Batista RB, Schultz N, Leite JM, Maia e Sá J, Barbosa KP, Resende AS, Alves BJR, Boddey RM (2012) Evidence from field nitrogen balance and 15N natural abundance data for the contribution of biological N2 fixation to Brazilian sugarcane varieties. Plant Soil 356:5–21

    Article  CAS  Google Scholar 

  • van Berkum P, Fuhrmann JJ (2000) Evolutionary relationships among the soybean bradyrhizobia reconstructed from 16S rRNA gene and internally transcribed spacer region sequence divergence. Int J Syst Evol Microbiol 50:2165–2172

    Article  PubMed  Google Scholar 

  • van Berkum P, Elia P, Song Q, Eardly BD (2012) Development and application of a multilocus sequence analysis method for the identification of genotypes within genus Bradyrhizobium and for establishing nodule occupancy of soybean (Glycine max L. Merr). Mol Plant Microbe Interact 25:321–330

    Article  PubMed  Google Scholar 

  • VanInsberghe D, Maas KR, Cardenas E, Strachan CR, Hallam SJ, Mohn WW (2015) Non-symbiotic Bradyrhizobium ecotypes dominate North American forest soils. The ISME journal 9:2435–2441

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vincent JM (1970) A manual for the practical study of the root-nodule bacteria. Blackwell Scientific, p 164

  • Vinuesa P, Silva C, Werner D, Martínez-Romero E (2005) Population genetics and phylogenetic inference in bacterial molecular systematics: the roles of migration and recombination in Bradyrhizobium species cohesion and delineation. Mol Phylogenet Evol 34:29–54

    Article  CAS  PubMed  Google Scholar 

  • Wang JY, Wang R, Zhang YM, Liu HC, Chen WF, Wang ET, Sui XH, Chen WX (2013) Bradyrhizobium daqingense sp. nov., isolated from soybean nodules. Int J Syst Evol Microbiol 63:616–624

    Article  CAS  PubMed  Google Scholar 

  • Willems A, Munive A, de Lajudie P, Gillis M (2003) In most Bradyrhizobium groups sequence comparison of 16S-23S rDNA internal transcribed spacer regions corroborates DNA-DNA hybridizations. Syst Appl Microbiol 26:203–210

    Article  CAS  PubMed  Google Scholar 

  • Yanni YG, Rizk RY, Corich V, Squartini A, Ninke K, Philip-Hollingsworth S, Orgambide G, de Bruijn F, Stoltzfus J, Buckley D, Schmidt TM, Mateos PF, Ladha JK, Dazzo FB (1997) Natural endophytic association between Rhizobium leguminosarum bv. trifolii and rice roots and assessment of its potential to promote rice growth. Plant Soil 194:99–114

    Article  CAS  Google Scholar 

  • Yao ZY, Kan FL, Wang ET, Wei GH, Chen WX (2002) Characterization of rhizobia that nodulate legume species of the genus Lespedeza and description of Bradyrhizobium yuanmingense sp. nov. Int J Syst Evol Microbiol 52:2219–2230

    CAS  PubMed  Google Scholar 

  • Yu X, Cloutier S, Tambong JT, Bromfield ES (2014) Bradyrhizobium ottawaense sp. nov., a symbiotic nitrogen fixing bacterium from root nodules of soybeans in Canada. Int J Syst Evol Microbiol 64:3202–3207

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang YM, Li Y Jr, Chen WF, Wang ET, Sui XH, Li QQ, Zhang YZ, Zhou YG, Chen WX (2012) Bradyrhizobium huanghuaihaiense sp. nov. an effective symbiotic bacterium isolated from soybean (Glycine max L.) nodules. Int J Syst Evol Microbiol 62:1951–1957

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors are grateful to Fernanda dos Santos Dourado, Natália Neutzling Camacho and Karine Moura de Freitas (Embrapa Agrobiologia) for technical support. Financial support by CNPq (projeto Universal process 477231/2012-8) and Faperj (processos E-26/111.398/2012 and E-26/010.001282/2015). The first author received a post-graduate fellowship from CNPq. The genome sequences of Bradyrhizobium daqingense (JGI Project ID: 1041378) and B. huanghuaihaiense (JGI Project ID: 1041383) were provided by a collaboration between the China General Microbiological Culture Collection Center and the US Department of Energy Joint Genome Institute, a DOE Office of Science User Facility, supported under Contract No. DE-AC02-05CH11231.

Author information

Authors and Affiliations

  1. Embrapa Agrobiologia, Rodovia BR 465 km 7, Seropédica, Rio De Janeiro, 23890-000, Brazil

    Gustavo Feitosa de Matos, Jerri Edson Zilli, Jean Luiz Simões de Araújo, José Ivo Baldani & Luc Felicianus Marie Rouws

  2. Universidade Federal Rural do Rio de Janeiro, Curso de Pós-graduação em Fitotecnia, Rodovia BR 465 km 07, Seropédica, Rio De Janeiro, 23890-000, Brazil

    Gustavo Feitosa de Matos

  3. Embrapa Meio Ambiente, C.P. 69, Jaguariúna, São Paulo, 13820-000, Brazil

    Marcia Maria Parma & Itamar Soares Melo

  4. Helmholtz Zentrum München, Research Unit Comparative Microbiome Analysis, Ingolstädter Landtraße 1, 85764, Oberschleißheim, Germany

    Viviane Radl

Authors
  1. Gustavo Feitosa de Matos
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jerri Edson Zilli
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jean Luiz Simões de Araújo
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Marcia Maria Parma
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Itamar Soares Melo
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Viviane Radl
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. José Ivo Baldani
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Luc Felicianus Marie Rouws
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Luc Felicianus Marie Rouws.

Additional information

Communicated by Erko Stackebrandt.

Electronic supplementary material

Below is the link to the electronic supplementary material.

203_2017_1398_MOESM1_ESM.pptx

Fig. S1 Maximum likelihood phylogram estimated from 583 nucleotide positions of the internally transcribed spacer (ITS) between the 16S and 23S rRNA genes. Bootstrap values are shown when the represented relationships were observed in at least 50% of 500 pseudoreplicates. The ML tree was obtained using the Kimura 2-parameter +G model. The scale bar represents the number of base pair substitutions per site. Genbank accession numbers are shown between parentheses. (PPTX 49 kb)

203_2017_1398_MOESM2_ESM.pptx

Fig. S2 Maximum likelihood phylogram estimated from 441 nucleotide positions of the nodC gene. Bootstrap values are shown when the represented relationships were observed in at least 50% of 500 pseudoreplicates. The ML tree was obtained using the Kimura 3-parameter +G +I model. The scale bar represents the number of base pair substitutions per site. Genbank accession numbers are shown between parentheses. (PPTX 55 kb)

Table S1 Genbank accession numbers of gene sequences used in this study. (DOCX 20 kb)

203_2017_1398_MOESM4_ESM.docx

Table S2 Percentage of nucleotide sequence similarity between gene sequences of Bradyrhizobium sacchari strain BR 10280T and other strains. (DOCX 20 kb)

Table S3 Carbon source usage features of B. sacchari strains and the related B. huanghuaihaiense. (DOCX 22 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

de Matos, G.F., Zilli, J.E., de Araújo, J.L.S. et al. Bradyrhizobium sacchari sp. nov., a legume nodulating bacterium isolated from sugarcane roots. Arch Microbiol 199, 1251–1258 (2017). https://doi.org/10.1007/s00203-017-1398-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00203-017-1398-6

Keywords

Search

Navigation