Plant and Soil

, Volume 287, Issue 1–2, pp 59–68 | Cite as

Screening for PGPR to improve growth of Cistus ladanifer seedlings for reforestation of degraded mediterranean ecosystems

  • Ramos Solano B.Email author
  • M. T. Pereyra de la Iglesia
  • A. Probanza
  • J. A. Lucas García
  • M. Megías
  • F. J. Gutierrez Mañero


A screening for PGPRs was carried out in the rhizosphere of wild populations of Cistus ladanifer. Two hundred and seventy bacteria were isolated, purified and grouped by morphological criteria. Fifty percent of the isolates were selected and tested for aminocyclopropanecarboxylic acid (ACC) degradation, auxin and siderophore production and phosphate solubilisation. Fifty-eight percent of the isolates showed at least one of the evaluated activities, with phosphate solubilisation and siderophore production being the most abundant traits. After PCR-RAPDs (Randomly amplified polymorphic DNA) analysis, 11 groups appeared with 85% similiarity, revealing the low diversity in the system. One strain of each group was tested in a biological assay, and those that enhanced Cistus growth were identified by 16S rDNA sequencing.

Although seven of the 11 assayed strains were phosphate solubilisers and able to produce siderophores, only one was really effective in increasing all biometric parameters in Cistus ladanifer seedlings, the lack of effect of the other six probably being due to the rich substrate used. This suggests that other mechanisms apart from nutrient mobilisation might be involved in growth promotion by this strain. However, the low diversity together with the high redundancy detected by PCR-RAPDs and the predominance of strains able to mobilise nutrients in the rhizosphere of Cistus reveals that the plant selects for bacteria that can help to supply scarce nutrients. This type of plant growth promoting rhizobacteria (PGPR) strains should be succesful in reforestation practices.


genetic diversity PGPR phosphate solubilisation rhizosphere reforestation 


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This study has been financed by project FEDER 1FD97-1441.We would like to thank Linda Hamalainen for editorial help and Mª Josefa Fernandez for technical support. M.T. Pereyra was formerly a Comunidad Autonoma de Madrid postdoctoral student.


  1. Alexander D B, Zuberer D A 1991 Use of chrome azurol S reagents to evaluate siderophore production by rhizosphere bacteria Biol. Fertil. Soils 12: 39–45CrossRefGoogle Scholar
  2. Benizri E, Courtade A, Picard C, Guckert A 1998 Role of maize root exudates in the production of auxins by Pseudomonas fluorescens M.3.1 Soil Biol. Biochem. 30: 1481–1484CrossRefGoogle Scholar
  3. Bowen G D, Rovira A D 1999 The rhizosphere and its management to improve plant growth Adv. Agron. 66: 1–103Google Scholar
  4. Burdman S, Jurkevith E, Okon Y 2000 Recent advances in the use of plant growth promoting rhizobacteria (PGPR) in agriculture Microb. Interact. Agric. Forest. 2: 229–250Google Scholar
  5. Cattelan A J, Hartel P G, Fuhrmann J J 1999 Screeening for plant growth-promoting rhizobacteria to promote early soybean growth Soil Sci. Soc. Am. J. 63: 1670–1680CrossRefGoogle Scholar
  6. Clark A G, Lanigan C M S 1993 Prospects for estimating nucleotide divergence with RAPDs Mol. Biol. Evol. 10: 1096–1111PubMedGoogle Scholar
  7. de Freitas J R, Banerjee M R, Germida J J 1997 Phosphate-solubilizing rhizobacteria enhance the growth and yield buy not phosphorus uptake of canola (Brassica napus L.) Biol. Fertil. Soils 24: 358–364CrossRefGoogle Scholar
  8. di Cello F, Bevivino A, Chiarini L, Fani R, Paffetti D, Tabacchioni S and Dalmastri C 1997 Biodiversity of a Burkholderia cepacia population isolated from the maize rhizosphere at different plant growth stages. Appl. Environ. Microbiol. 63, 4485–4493Google Scholar
  9. Enebak S A, Wei G, Kloepper J W 1997 Effects of plant growth-promoting rhizobacteria on loblolly and slash pine seedlings Forest Sci. 44: 139–144Google Scholar
  10. Glick B R, Karaturovic D M, Newell P C 1995 A novel procedure for rapid isolation of plant growth promoting pseudomonads Can. J. Microbiol. 41: 533–536CrossRefGoogle Scholar
  11. Goddard V J, Bailey M J, Darrah P, Lilley A K, Thompson I P 2001 Monitoring temporal and spatial variation in rhizosphere bacterial population diversity: A community approach for the improved selection of rhizosphere competent bacteria Plant Soil 232: 181–193CrossRefGoogle Scholar
  12. Gonzalez-Lopez J, Salmeron V, Martinez-Toledo M V, Ballesteros F, Ramos-Cormenzana A 1986 Production of auxins, gibberellins and cytokinins by Azotobacter vinelandii ATTCC12837 in chemically-defined media and dialyses soil media Soil Biol. Biochem. 18: 119–120CrossRefGoogle Scholar
  13. Gutierrez Mañero F J, Acero N, Lucas J A, Probanza A 1996 The influence of native rhizobacteria on European alder [Alnus glutinosa (L.) Gaertn.] growth. II. Characterization of growth promoting and growth inhibiting strains Plant Soil 182: 67–74CrossRefGoogle Scholar
  14. Gutierrez Mañero F J, Ramos B, Lucas García J A, Probanza A, Barrientos M L 2002 Systemic induction of terpenic compounds in D. lanata J. Plant Physiol. 160: 105–113CrossRefGoogle Scholar
  15. Gutierrez Mañero F J, Ramos Solano B, Probanza A, Mehouachi J, Tadeo F R, Talon M 2001 The plant growth-promoting rhizobacteria Bacillus pumilus and Bacillus licheniformis produce high amounts of physiologically active gibberellins Physiol. Plant. 111: 1–7CrossRefGoogle Scholar
  16. Harman J H 1967 Modern Factor Analysis. 2nd ed. Univ. Chicago Press, ChicagoGoogle Scholar
  17. Kloepper J W, Scrhoth M N, Miller T D 1980 Effects of rhizosphere colonization by plant growth-promoting rhizobacteria on potato plant development and yield Phytopathology 70: 1078–1082CrossRefGoogle Scholar
  18. Kucey R M N 1983 Phosphate-solubilizing bacteria and fungi in various cultivated and virgin Alberta soils Can. J. Soil Sci. 63: 671–678Google Scholar
  19. Louws F J, Rademaker J L W, de Bruijn F J 1999 The three Ds of PCR-based genomic analysis of phytobacteria: Diversity, detection and disease diagnosis Annu. Rev. Phytpathol. 37: 81–125CrossRefGoogle Scholar
  20. Lucas García J A, Probanza A, Ramos B, Gutierrez Mañero F J 2001 Genetic variability of rhizobacteria from wild populations of four Lupinus species based on PCR-RAPDs J. Plant Nutr. Soil Sci. 164: 1–7CrossRefGoogle Scholar
  21. Lynch J M 1990 The Rhizosphere. Wiley-Interscience, Chichester, EnglandGoogle Scholar
  22. Marilley L, Aragno M 1999 Phylogenetic diversity of bacterial communities differing in degree of proximity of Lolium perenne and Trifolium repens roots Appl. Soil Ecol. 13: 127–136CrossRefGoogle Scholar
  23. Marten P, Brueckner S, Berg G 2001 Biological plant protection using rhizobacteria – an environmental friendly alternative for biologicl control of soilborne and seedborne phytopathogenic fungi Gesunde Pflanzen 53: 224–234Google Scholar
  24. Nei M, Miller J C 1990 A simple method for estimating average number of nucleotide substitutons within and between populations from restriction data Genetics 125: 873–879PubMedGoogle Scholar
  25. Probanza A, Mateos J L, Lucas J A, Ramos B, de Felipe M R, Gutierrez Mañero F J 2001 Effects of inoculation with PGPR Bacillus and Pisolitus tinctorius on Pinus pinea L. growth, bacterial rhizosphere colonization and mycorrhizal infection Microbial Ecol. 41: 140–148Google Scholar
  26. Rainey P B 1999 Adaptation of Pseudomonas fluorescens to the plant rhizosphere Environ. Microbiol. 1: 243–257PubMedCrossRefGoogle Scholar
  27. Ramamoorthy V, Viswanathan R, Raguchander T, Prakasam V, Samiyappan R 2001 Induction of systemic resistance by plant growth promoting rhizobacteria in crop plants against pests and diseases Crop Protect. 20: 1–11CrossRefGoogle Scholar
  28. Ramos B, Lucas García J A, Probanza A, Barrientos M L, Gutierrez Mañero F J 2002 Alterations in the rhizobacterial community associated with European alder growth when inoculated with PGPR strain Bacillus licheniformis Environ. Exp. Bot. 49: 61–68CrossRefGoogle Scholar
  29. Richardson A E 2001 Prospects for using soil microorganism to improve the acquisition of phosphorus by plants Aust. J. Plant Physiol. 28: 897–906Google Scholar
  30. Rodriguez H, Rossolini G M, Gonzalez T, Li J, Glick B R 2000 Isolation of a gene from Burkholderia cepacia IS16 encoding a protein that facilitates phosphatase activity Curr. Microbiol. 40: 362–366PubMedCrossRefGoogle Scholar
  31. Selvadurai E L, Brown A E, Hamilton J T G 1991 Production of indole-3-acetic acid analogues by strains of Bacillus cereus in relation to their influence on seedling development Soil Biol. Biochem. 23: 401–403CrossRefGoogle Scholar
  32. Van Loon L C, Bakker P A H M, Pieterse C M J 1998 Systemic resistance induced by rhizosphere bacteria Annu. Rev. Phytopathol. 36: 453–483PubMedCrossRefGoogle Scholar
  33. Wayne L G, Brenner D J, Colwell R R, Grimont P A D, Kandler O, Kirchevsdy M I, Moore L H, Moore W E C, Murray R G E, Stackerbrandt E, Starr M P, Turper H G 1987 Report of the ad hoc committee on reconciliation of approaches to bacterial systematics Int. J. Syst. Bacteriol. 37: 463–464CrossRefGoogle Scholar
  34. Wiehe W, Höflich G 1995 Establishment of plant growth promoting bacteria in the rhizosphere of subsequent plants after harvest of the inoculated precrops Microbiol. Res. 150: 331–336Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2006

Authors and Affiliations

  • Ramos Solano B.
    • 1
    Email author
  • M. T. Pereyra de la Iglesia
    • 1
  • A. Probanza
    • 1
  • J. A. Lucas García
    • 1
  • M. Megías
    • 2
  • F. J. Gutierrez Mañero
    • 1
  1. 1.Facultad FarmaciaUniversidad San Pablo CEUMadridSpain
  2. 2.Fac. FarmaciaUniversidad de SevillaSevillaSpain

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