Skip to main content
SpringerLink
Log in

Effects of N-starvation and C-source on Bradyrhizobium japonicum exopolysaccharide production and composition, and bacterial infectivity to soybean roots

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

Abstract

The exopolysaccharide (EPS) is an extracellular molecule that in Bradyrhizobium japonicum affects bacterial efficiency to nodulate soybean. Culture conditions such as N availability, type of C-source, or culture age can modify the amount and composition of EPS. To better understand the relationship among these conditions for EPS production, we analyzed their influence on EPS in B. japonicum USDA 110 and its derived mutant ΔP22. This mutant has a deletion including the 3′ region of exoP, exoT, and the 5′ region of exoB, and produces a shorter EPS devoid of galactose. The studies were carried out in minimal media with the N-source at starving or sufficient levels, and mannitol or malate as the only C-source. Under N-starvation there was a net EPS accumulation, the levels being similar in the wild type and the mutant with malate as the C-source. By contrast, the amount of EPS diminished in N-sufficient conditions, being poyhydroxybutyrate accumulated with culture age. Hexoses composition was the same in both N-situations, either with mannitol or malate as the only C-source, in contrast to previous observations made with different strains. This result suggests that the change in EPS composition in response to the environment is not general in B. japonicum. The wild type EPS composition was 1 glucose:0.5 galactose:0.5 galacturonic acid:0.17 mannose. In ΔP22 the EPS had no galactose but had galacturonic acid, thus indicating that it was not produced from oxidation of UDP-galactose. Infectivity was lower in ΔP22 than in USDA 110. When the mutant infectivity was compared between N-starved or N-sufficient cultures, the N-starved were not less infective, despite the fact that the amounts of altered EPS produced by this mutant under N-starvation were higher than in N-sufficiency. Since this altered EPS does not bind soybean lectin, the interaction of EPS with this protein was not involved in increasing ΔP22 infectivity under N-starvation.

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

Abbreviations

CPS:

Capsular polysaccharide

DAI:

Days after inoculation

EPS:

Exopolysaccharide

Gal:

Galactose

GalA:

Galacturonic acid

Glc:

Glucose

GlcA:

Glucuronic acid

Mal:

Malate

Man:

Mannose

Mtl:

Mannitol

PHB:

Polyhydroxybutyrate

OD500 :

Optical density at 500 nm

RDU:

Relative distance unit

RT:

Root tip

SBL:

Soybean lectin

SERH:

Smallest emergent root hairs

References

  • Amarasingham CR, Davis BD (1965) Regulation of α-ketoglutarate dehydrogenase formation in Escherichia coli. J Biol Chem 240:3664–3668

    PubMed  CAS  Google Scholar 

  • An J, Carlson RW, Glushka J, Streeter JG (1995) The structure of a novel polysaccharide produced by Bradyrhizobium species within soybean nodules. Carbohydr Res 269:303–317

    Article  PubMed  CAS  Google Scholar 

  • Becker BU, Kosch K, Parniske M, Müller P (1998) Exopolysaccharide (EPS) synthesis in Bradyrhizobium japonicum: sequence, operon structure and mutational analysis of an exo gene cluster. Mol Gen Genet 259:161–171

    Article  PubMed  CAS  Google Scholar 

  • Bhuvaneswari TV, Turgeon BG, Bauer WD (1980) Early events in the infection of soybean (Glycine max L. Merr) by Rhizobium japonicum. I. Localization of infectible root cells. Plant Physiol 66:1027–1031

    PubMed  Google Scholar 

  • Bhuvaneswari BV, Mills KK, Crist DK, Evans WR, Bauer WD (1983) Effects of culture age on symbiotic infectivity of Rhizobium japonicum. J Bacteriol 153:443–451

    PubMed  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  PubMed  CAS  Google Scholar 

  • Cevallos M, Encarnación S, Leija A, Mora Y, Mora J (1996) Genetic and physiological characterization of a Rhizobium etli mutant strain unable to synthesize poly-beta-hydroxybutyrate. J Bacteriol 178:1646–1654

    PubMed  CAS  Google Scholar 

  • Encarnación S, Dunn M, Willms K, Mora J (1995) Fermentative and aerobic metabolism in Rhizobium etli. J Bacteriol 177:3058–3066

    PubMed  Google Scholar 

  • Encarnación S, Vargas MC, Dunn MF, Dávalos A, Mendoza G, Mora Y, Mora J (2002) AniA regulates reserve polymer accumulation and global protein expression in Rhizobium etli. J Bacteriol 184:2287–2295

    Article  PubMed  Google Scholar 

  • Fraysse N, Couderc F, Poinsot V (2003) Surface polysaccharide involvement in establishing the Rhizobium-legume symbiosis. Eur J Biochem 270:1365–1380

    Article  PubMed  CAS  Google Scholar 

  • Gage DJ (2004) Infection and invasion of roots by symbiotic, nitrogen-fixing rhizobia during nodulation of temperate legumes. Microbiol Mol Biol Rev 68:280–300

    Article  PubMed  CAS  Google Scholar 

  • González JE, Semino CE, Wang L-X, Castellano-Torres LE Walker GC (1998) Biosynthetic control of molecular weight in the polymerization of the octasaccharide subunits of succinoglycan, a symbiotically important exopolysaccharide of Rhizobium meliloti. Proc Natl Acad Sci USA 95:13477–13482

    Article  PubMed  Google Scholar 

  • Götz R, Limmer N, Ober K, Schmitt R (1982) Motility and chemotaxis in two strains of Rhizobium with complex flagella. J Gen Microbiol 128:789–798

    Google Scholar 

  • Hozbor DF, Pich Otero AJL, Lodeiro AR, Del Papa MF, Pistorio M, Lagares A (2004) Functional complementation among different surface polysaccharides in the symbiosis between Sinorhizobium meliloti and Medicago species. Res Microbiol 155:855–860

    Article  PubMed  CAS  Google Scholar 

  • Kaneko T, Nakamura Y, Sato S et al (2002) Complete genomic sequence of nitrogen-fixing symbiotic bacterium Bradyrhizobium japonicum USDA 110. DNA Res 9:189–197

    Article  PubMed  Google Scholar 

  • Karr DB, Liang R-T, Reuhs BL, Emerich DW (2000) Altered exopolysaccharides of Bradyrhizobium japonicum mutants correlate with impaired soybean lectin binding, but not with effective nodule formation. Planta 211:218–226

    Article  PubMed  CAS  Google Scholar 

  • Kereszt A, Kiss E, Reuhs BL, Carlson RW, Kondorosi A, Putnoky P (1998) Novel rkp gene clusters of Sinorhizobium meliloti involved in capsular polysaccharide production and invasion of the symbiotic nodule: the rkpK gene encodes a UDP-glucose dehydrogenase. J Bacteriol 180:5426–5431

    PubMed  CAS  Google Scholar 

  • Laus MC, Logman TJ, Van Brussel AA, Carlson RW, Azadi P, Gao MY, Kijne JW (2004) Involvement of exo5 in production of surface polysaccharides in Rhizobium leguminosarum and its role in nodulation of Vicia sativa subsp. nigra. J Bacteriol 186:6617–6625

    Article  PubMed  CAS  Google Scholar 

  • Law JH, Slepecky RA (1961) Assay of poly-hydroxybutyric acid. J Bacteriol 82:33–36

    PubMed  CAS  Google Scholar 

  • Lis H, Sela BA, Sachs L, Sharon N (1970) Specific inhibition by N-acetyl-d-galactosamine of the interaction between soybean agglutinin and animal cell surfaces. Biochim Biophys Acta 211:582–585

    Article  PubMed  CAS  Google Scholar 

  • Lodeiro AR, González P, Hernández A, Balagué LJ, Favelukes G (2000) Comparison of drought tolerance in nitrogen-fixing and inorganic nitrogen-grown common beans. Plant Sci 154:31–41

    Article  PubMed  CAS  Google Scholar 

  • López-García S, Vázquez TE E, Favelukes G, Lodeiro A (2001) Improved soybean root association of N-starved Bradyrhizobium japonicum. J Bacteriol 183:7241–7252

    Article  PubMed  CAS  Google Scholar 

  • Louch HA, Miller KJ (2001) Synthesis of a low-molecular-weight form of exopolysaccharide by Bradyrhizobium japonicum USDA 110. Appl Environ Microbiol 67:1011–1014

    Article  PubMed  CAS  Google Scholar 

  • Mort AJ, Bauer WD (1980) Composition of the capsular and extracellular polysaccharides of Rhizobium japonicum. Changes with culture age and correlations with binding of soybean seed lectin to the bacteria. Plant Physiol 66:158–163

    Article  PubMed  CAS  Google Scholar 

  • Mort AJ, Bauer WD (1982) Application of two new methods for cleavage of polysaccharides into specific oligosaccharide fragments. Structure of the capsular and extracellular polysaccharides of Rhizobium japonicum that bind soybean lectin. J Biol Chem 257:1870–1875

    PubMed  CAS  Google Scholar 

  • Niehaus K, Baier R, Kohring B, Flashl E, Pühler A (1997) Symbiotic suppression of the Medicago sativa plant defence system by Rhizobium meliloti oligosaccharides. In: Legoki A, Bothe H, Pühler A (eds) Biological fixation of nitrogen for ecology and sustainable agriculture. Springer, Berlin Heidelberg New York, pp 110–114

    Google Scholar 

  • Niemeyer D, Becker A (2001) The molecular weight distribution of succinoglycan produced by Sinorhizobium meliloti is influenced by specific tyrosine phosphorylation and ATPase activity of the cytoplasmic domain of the ExoP protein. J Bacteriol 183:5163–5170

    Article  PubMed  CAS  Google Scholar 

  • Ong LC, Lin Y-H (2003) Metabolite profiles and growth characteristics of Rhizobium meliloti cultivated at different specific growth rates. Biotechnol Prog 19:714–719

    Article  PubMed  CAS  Google Scholar 

  • Panzieri M, Marchettini N, Hallam TG (2000) Importance of the Bradyrhizobium japonicum symbiosis for the sustainability of a soybean cultivation. Ecol Model 135:301–310

    Article  Google Scholar 

  • Parniske M, Kosch K, Werner D, Müller P (1993) ExoB mutants of Bradyrhizobium japonicum with reduced competitiveness for nodulation of Glycine max. Mol Plant-Microbe Interact 6:99–106

    CAS  Google Scholar 

  • Parniske M, Schmidt PE, Kosch K, Müller P (1994) Plant defense responses of host plants with determinate nodules induced by EPS-defective exoB mutants of Bradyrhizobium japonicum. Mol Plant-Microbe Interact 7:631–638

    CAS  Google Scholar 

  • Patriarca EJ, Tatè R, Ferraioli S, Iaccarino M (2004) Organogenesis of the legume root nodules. Int Rev Cytol 234:201–262

    Article  PubMed  Google Scholar 

  • Streeter JG, Salminen SO, Whitmoyer RE, Carlson RW (1992) Formation of novel polysaccharides by Bradyrhizobium japonicum bacteroids in soybean nodules. Appl Environ Microbiol 58:607–613

    PubMed  CAS  Google Scholar 

  • Trevelyan WE, Harrison JS (1952) Studies on yeast metabolism. I. Fractionation and microdetermination of cell carbohydrates. Biochem J 50:298–310

    PubMed  CAS  Google Scholar 

  • van Rhijn P, Goldberg RB, Hirsch AM (1998) Lotus corniculatus nodulation specificity is changed by the presence of a soybean lectin gene. Plant Cell 10:1233–1249

    Article  PubMed  Google Scholar 

  • Vincent J M (1970) A manual for the practical study of the root nodule bacteria. IBP handbook No. 15. Blackwell, Oxford

    Google Scholar 

Download references

Acknowledgments

This work was supported by International Foundation for Science (Sweden) grant No. C/2736-2 and Agencia Nacional de Promoción Científica y Tecnológica grant No. PICT 08-9809. JIQ and EJM are Doctoral fellows of CONICET; SLLG is a Postdoctoral fellow of CONICET; MJA and JPG are Doctoral fellows of ANPCyT; AC and ARL are members of the Scientific Career of CONICET, Argentina. The authors are grateful to A. McCarthy and N. Stern for English revision.

Author information

Authors and Affiliations

  1. Instituto de Bioquímica y Biología Molecular (IBBM), Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, Calles 47 y 115, 1900, La Plata, Argentina

    Juan Ignacio Quelas, Silvina L. López-García, M. Julia Althabegoiti, Elías J. Mongiardini, Julieta Pérez-Giménez & Aníbal R. Lodeiro

  2. Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR), Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón II, 3er Piso. Ciudad Universitaria, 1428, Buenos Aires, Argentina

    Adriana Casabuono & Alicia Couto

Authors
  1. Juan Ignacio Quelas
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Silvina L. López-García
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Adriana Casabuono
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. M. Julia Althabegoiti
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Elías J. Mongiardini
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Julieta Pérez-Giménez
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Alicia Couto
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Aníbal R. Lodeiro
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Aníbal R. Lodeiro.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Quelas, J.I., López-García, S.L., Casabuono, A. et al. Effects of N-starvation and C-source on Bradyrhizobium japonicum exopolysaccharide production and composition, and bacterial infectivity to soybean roots. Arch Microbiol 186, 119–128 (2006). https://doi.org/10.1007/s00203-006-0127-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00203-006-0127-3

Keywords

Search

Navigation