Microbial Ecology

, Volume 78, Issue 3, pp 699–713 | Cite as

Comparative Genomics, Siderophore Production, and Iron Scavenging Potential of Root Zone Soil Bacteria Isolated from ‘Concord’ Grape Vineyards

  • Ricky W. Lewis
  • Anjuman Islam
  • Lee Opdahl
  • Joan R. Davenport
  • Tarah S. SullivanEmail author
Plant Microbe Interactions


Iron (Fe) deficiency in crop production is a worldwide problem which often results in chlorosis in grapevines, particularly in calcareous soils. Siderophores secreted by microorganisms and Strategy II plants can chelate Fe and other metals in soil solution, and siderophore-Fe complexes can then be utilized by plants and microbes. Plants may also shift rhizosphere conditions to favor siderophore-producing microbes, which can increase plant available Fe. Between-row cover crops (barley, rye, wheat, wheat/vetch) were planted as living mulch to address grapevine chlorosis by enhancing soil health in two vineyards in central Washington. The objectives of the current study were to (1) enrich for siderophore-producing organisms from within the indigenous rooting zone community of ‘Concord’ grapevines, and (2) perform comparative genomics on putative siderophore producing organisms to assess potentially important Fe acquisition-related functional domains and protein families. A high-throughput, chrome azurol S (CAS)-based enrichment assay was used to select siderophore-producing microbes from ‘Concord’ grapevine root zone soil. Next-generation whole genome sequencing allowed the assembly and annotation of ten full genomes. Phylogenetic analysis revealed two distinct clades among the genomes using the 40 nearest neighbors available in the public database, all of which were of the Pseudomonas genus. Significant differences in functional domain abundances were observed between the clades including iron acquisition and metabolism of amino acids, carbon, nitrogen, phosphate, and sulfur. Diverse mechanisms of Fe uptake and siderophore production/uptake were identified in the protein families of the genomes. The sequenced organisms are likely pseudomonads which are well-suited for iron scavenging, suggesting a potential role in Fe turnover in vineyard systems.


Rhizosphere function Chrome azurol S (CAS) enrichment Microbial cheating Grapevine microbiome Pseudomonas genomics Grapevine nutrition 



The authors wish to thank Kalyani Muhunthan for assistance in laboratory procedures and the members of the Davenport Lab for assistance with sampling and plant tissue analyses.

Author Contributions

Ricky W. Lewis performed whole genome sequencing, processed and analyzed the sequencing data, and assisted with manuscript writing. Anjuman Islam gathered samples, performed the microplate siderophore production assay, extracted DNA, and assisted with manuscript writing. Lee Opdahl assisted with whole genome sequencing, interpretation of results, and manuscript writing. Tarah S. Sullivan and Joan R. Davenport conceived of the experimental design and assisted with sample gathering, interpretation of results, and manuscript writing.

Funding Information

Funding was provided by the Washington State Concord Grape Research Council, by the Washington State University BioAg program, and by the USDA/NIFA through Hatch project 1014527.

Compliance with Ethical Standards

Competing Interests

The authors declare that they have no competing interests.

Supplementary material

248_2019_1324_MOESM1_ESM.docx (16 kb)
ESM 1 (DOCX 15.6 kb)
248_2019_1324_MOESM2_ESM.docx (20 kb)
ESM 2 (DOCX 20.4 kb)


  1. 1.
    Davenport JR, Stevens RG (2006) High soil moisture and low soil temperature are associated with chlorosis occurrence in Concord grape. Hortscience 41:418–422CrossRefGoogle Scholar
  2. 2.
    Pradubsuk S, Davenport JR (2010) Seasonal uptake and partitioning of macronutrients in mature ‘Concord’ grape. J Am Soc Hortic Sci 135:474–483CrossRefGoogle Scholar
  3. 3.
    Abadia J, Vazquez S, Rellan-Alvarez R, El-Jendoubi H, Abadia A, Alvarez-Fernandez A, Lopez-Millan AF (2011) Towards a knowledge-based correction of iron chlorosis. Plant Physiol Biochem 49:471–482. CrossRefGoogle Scholar
  4. 4.
    Garcia-Mina JM, Bacaicoa E, Fuentes M, Casanova E (2013) Fine regulation of leaf iron use efficiency and iron root uptake under limited iron bioavailability. Plant Sci 198:39–45. CrossRefGoogle Scholar
  5. 5.
    Singh K, Singh Y, Upadhyay AK, Mori S (2011) Phytosiderophore-based molecular approach for enhanced iron acquisition to increase crop production under high pH calcareous soils. Indian J Agric Sci 81:679–689Google Scholar
  6. 6.
    Colombo C, Palumbo G, He JZ, Pinton R, Cesco S (2014) Review on iron availability in soil: interaction of Fe minerals, plants, and microbes. J Soils Sediments 14:538–548. CrossRefGoogle Scholar
  7. 7.
    Kim SA, Guerinot ML (2007) Mining iron: iron uptake and transport in plants. FEBS Lett 581:2273–2280. CrossRefGoogle Scholar
  8. 8.
    Lopez-Millan AF, Grusak MA, Abadia A, Abadia J (2013) Iron deficiency in plants: an insight from proteomic approaches. Front Plant Sci 4.
  9. 9.
    Rajaie M, Tavakoly AR (2018) Iron and/or acid foliar spray versus soil application of Fe-EDDHA for prevention of iron deficiency in Valencia orange grown on a calcareous soil. J Plant Nutr 41:150–158. Google Scholar
  10. 10.
    Bavaresco L, Goncalves M, Civardi S, Gatti M, Ferrari F (2010) Effects of traditional and new methods on overcoming lime-induced chlorosis of grapevine. Am J Enol Vitic 61:186–190Google Scholar
  11. 11.
    Gamble AV, Howe JA, Delaney D, van Santen E, Yates R (2014) Iron chelates alleviate iron chlorosis in soybean on high pH soils. Agron J 106:1251–1257. CrossRefGoogle Scholar
  12. 12.
    Smith BR, Cheng LL (2006) Fe-EDDHA alleviates chlorosis in ‘Concord’ grapevines grown at high pH. Hortscience 41:1498–1501CrossRefGoogle Scholar
  13. 13.
    Oburger E, Gruber B, Schindlegger Y, Schenkeveld WDC, Hann S, Kraemer SM, Wenzel WW, Puschenreiter M (2014) Root exudation of phytosiderophores from soil-grown wheat. New Phytol 203:1161–1174. CrossRefGoogle Scholar
  14. 14.
    Puschenreiter M, Gruber B, Wenzel WW, Schindlegger Y, Hann S, Spangl B, Schenkeveld WDC, Kraemer SM, Oburger E (2017) Phytosiderophore-induced mobilization and uptake of Cd, Cu, Fe, Ni, Pb and Zn by wheat plants grown on metal-enriched soils. Environ Exp Bot 138:67–76. CrossRefGoogle Scholar
  15. 15.
    Saha M, Sarkar S, Sarkar B, Sharma BK, Bhattacharjee S, Tribedi P (2016) Microbial siderophores and their potential applications: a review. Environ Sci Pollut Res 23:3984–3999. CrossRefGoogle Scholar
  16. 16.
    Jin CW, Li GX, Yu XH, Zheng SJ (2010) Plant Fe status affects the composition of siderophore-secreting microbes in the rhizosphere. Ann Bot 105:835–841. CrossRefGoogle Scholar
  17. 17.
    Masalha J, Kosegarten H, Elmaci O, Mengel K (2000) The central role of microbial activity for iron acquisition in maize and sunflower. Biol Fertil Soils 30:433–439CrossRefGoogle Scholar
  18. 18.
    Desai A, Archana G (2011) Role of siderophores in crop improvementGoogle Scholar
  19. 19.
    Ahmed E, Holmstrom SJM (2014) Siderophores in environmental research: roles and applications. Microb Biotechnol 7:196–208. CrossRefGoogle Scholar
  20. 20.
    Ahmed E, Holmstrom SJM (2015) Siderophore production by microorganisms isolated from a podzol soil profile. Geomicrobiol J 32:397–411. CrossRefGoogle Scholar
  21. 21.
    Crits-Christoph A, Diamond S, Butterfield CN, Thomas BC, Banfield JF (2018) Novel soil bacteria possess diverse genes for secondary metabolite biosynthesis. Nature 558:440–444. CrossRefGoogle Scholar
  22. 22.
    Barber MF, Eldel NC (2015) Buried treasure: evolutionary perspectives on microbial iron piracy. Trends Genet 31:627–636. CrossRefGoogle Scholar
  23. 23.
    Popat R, Harrison F, da Silva AC, Easton SAS, McNally L, Williams P, Diggle SP (2017) Environmental modification via a quorum sensing molecule influences the social landscape of siderophore production. Proc R Soc B Biol Sci 284:20170200. CrossRefGoogle Scholar
  24. 24.
    Castro RO, Bucio JL (2013) Small molecules involved in transkingdom communication between plants and rhizobacteria. In: de Bruijn FJ (ed) Molecular microbial ecology of the rhizosphere. Wiley, Hoboken, pp 295–307CrossRefGoogle Scholar
  25. 25.
    Butaite E, Baumgartner M, Wyder S, Kummerli R (2017) Siderophore cheating and cheating resistance shape competition for iron in soil and freshwater Pseudomonas communities. Nat Commun 8:414. CrossRefGoogle Scholar
  26. 26.
    O’Brien S, Lujan AM, Paterson S, Cant MA, Buckling A (2017) Adaptation to public goods cheats in Pseudomonas aeruginosa. Proc R Soc B Biol Sci 284:20171089. CrossRefGoogle Scholar
  27. 27.
    Lewis RW, LeTourneau MK, Davenport JR, Sullivan TS (2018) ‘Concord’ grapevine nutritional status and chlorosis rank associated with fungal and bacterial root zone microbiomes. Plant Physiol Biochem 129:429–436. CrossRefGoogle Scholar
  28. 28.
    Davenport JR, Stevens RG, Whitley KM (2008) Spatial and temporal distribution of soil moisture in drip-irrigated vineyards. Hortscience 43:229–235CrossRefGoogle Scholar
  29. 29.
    Singer SD, Davenport JR, Hoheisel G-A, Moyer M (2018) Vineyard nutrient management in Washington State.Google Scholar
  30. 30.
    Bremner J (1996) Nitrogen-total Methods of Soil Analysis Part 3—Chemical Methods: 1085–1121Google Scholar
  31. 31.
    Schwyn B, Neilands JB (1987) Universal chemical assay for the detection and determination of siderophores. Anal Biochem 160:47–56. CrossRefGoogle Scholar
  32. 32.
    Allen B, Drake M, Harris N, Sullivan T (2017) Using KBase to assemble and annotate prokaryotic genomes. Curr Protoc Microbiol 46:1E.13.1–1E.13.18. CrossRefGoogle Scholar
  33. 33.
    Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30:2114–2120. CrossRefGoogle Scholar
  34. 34.
    Nurk S, Bankevich A, Antipov D, Gurevich AA, Korobeynikov A, Lapidus A, Prjibelski AD, Pyshkin A, Sirotkin A, Sirotkin Y, Stepanauskas R, Clingenpeel SR, Woyke T, McLean JS, Lasken R, Tesler G, Alekseyev MA, Pevzner PA (2013) Assembling single-cell genomes and mini-metagenomes from chimeric MDA products. J Comput Biol 20:714–737. CrossRefGoogle Scholar
  35. 35.
    Wu Y-W, Simmons BA, Singer SW (2015) MaxBin 2.0: an automated binning algorithm to recover genomes from multiple metagenomic datasets. Bioinformatics 32:605–607CrossRefGoogle Scholar
  36. 36.
    Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, Tyson GW (2015) CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome research: gr. 186072.186114.Google Scholar
  37. 37.
    Gurevich A, Saveliev V, Vyahhi N, Tesler G (2013) QUAST: quality assessment tool for genome assemblies. Bioinformatics 29:1072–1075CrossRefGoogle Scholar
  38. 38.
    Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M (2013) The SEED and the rapid annotation of microbial genomes using subsystems technology (RAST). Nucleic Acids Res 42:D206–D214CrossRefGoogle Scholar
  39. 39.
    Price MN, Dehal PS, Arkin AP (2010) FastTree 2 – approximately maximum-likelihood trees for large alignments. PLoS One 5:e9490. CrossRefGoogle Scholar
  40. 40.
    Perez-Miranda S, Cabirol N, George-Tellez R, Zamudio-Rivera L, Fernandez F (2007) O-CAS, a fast and universal method for siderophore detection. J Microbiol Methods 70:127–131CrossRefGoogle Scholar
  41. 41.
    Sullivan TS, Ramkissoon S, Garrison VH, Ramsubhag A, Thies JE (2012) Siderophore production of African dust microorganisms over Trinidad and Tobago. Aerobiologia 28:391–401. CrossRefGoogle Scholar
  42. 42.
    Aznar A, Dellagi A (2015) New insights into the role of siderophores as triggers of plant immunity: what can we learn from animals? J Exp Bot 66:3001–3010. CrossRefGoogle Scholar
  43. 43.
    Lamont IL, Martin LW (2003) Identification and characterization of novel pyoverdine synthesis genes in Pseudomonas aeruginosa. Microbiology 149:833–842CrossRefGoogle Scholar
  44. 44.
    Moon CD, Zhang XH, Matthijs S, Schafer M, Budzikiewicz H, Rainey PB (2008) Genomic, genetic and structural analysis of pyoverdine-mediated iron acquisition in the plant growth-promoting bacterium Pseudomonas fluorescens SBW25. BMC Microbiol 8:7CrossRefGoogle Scholar
  45. 45.
    Meyer JM, Abdallah MA (1978) The fluorescent pigment of Pseudomonas fluorescens: biosynthesis, purification and physico-chemical properties. J Gen Microbiol 107:319–328CrossRefGoogle Scholar
  46. 46.
    Hohnadel D, Meyer JM (1988) Specificity of pyoverdine-mediated iron uptake among fluorescent Pseudomonas strains. J Bacteriol 170:4865–4873CrossRefGoogle Scholar
  47. 47.
    Mirleau P, Delmorme S, Philippot L, Meyer JM, Mazurier S, Lemanceau P (2000) Fitness in soil and rhizosphere of Pseudomonas fluorescens C7R12 compared with a C7R12 mutant affected in pyoverdine synthesis and uptake. FEMS Microbiol Ecol 34:35–44CrossRefGoogle Scholar
  48. 48.
    Cunrath O, Geoffroy VA, Schalk IJ (2016) Metallome of Pseudomonas aeruginosa: a role for siderophores. Environ Microbiol 18:3258–3267. CrossRefGoogle Scholar
  49. 49.
    Baldi F, Gallo M, Battistel D, Barbaro E, Gambaro A, Daniele S (2016) A broad mercury resistant strain of Pseudomonas putida secretes pyoverdine under limited iron conditions and high mercury concentrations. Biometals 29:1097–1106. CrossRefGoogle Scholar
  50. 50.
    O'Brien S, Hesse E, Lujan A, Hodgson DJ, Gardner A, Buckling A (2018) No effect of intraspecific relatedness on public goods cooperation in a complex community. Evolution 72:1165–1173. CrossRefGoogle Scholar
  51. 51.
    Buyer JS, DeLorenzo V, Neilands JB (1991) Production of the siderophore aerobactin by a halophilic pseudomonad. Appl Environ Microbiol 57:2246–2250Google Scholar
  52. 52.
    Sandy M, Butler A (2009) Microbial iron acquisition: marine and terrestrial siderophores. Chem Rev 109:4580–4595CrossRefGoogle Scholar
  53. 53.
    Valvano MA, Silver RP, Crosa JH (1986) Occurrence of chromosome- or plasmid-mediated aerobactin iron transport systems and hemolysin production among clonal groups of human invasive strains of Escherichia coli K1. Infect Immun 52:192–199Google Scholar
  54. 54.
    Fischbach MA, Lin H, Liu DR, Walsh CT (2006) How pathogenic bacteria evade mammalian sabotage in the battle for iron. Nat Chem Biol 2:132–138CrossRefGoogle Scholar
  55. 55.
    Chen LM, Dick WA, Streeter JG (2000) Production of aerobactin by microorganisms from a compost enrichment culture and soybean utilization. J Plant Nutr 23:2047–2060. CrossRefGoogle Scholar
  56. 56.
    Thode SK, Rojek E, Kozlowski M, Ahmad R, Haugen P (2018) Distribution of siderophore gene systems on a Vibrionaceae phylogeny: database searches, phylogenetic analyses and evolutionary perspectives. PLoS One 13:e0191860. CrossRefGoogle Scholar
  57. 57.
    Dertz EA, Raymond KN (2003) Siderophores and transferrins. In: Que L, Tolman WB (eds) Comprehensive coordination chemistry II. Elsevier, Ltd., PhiladelphiaGoogle Scholar
  58. 58.
    Miethke M, Marahiel MA (2007) Siderophore-based iron acquisition and pathogen control. Microbiol Mol Biol Rev 71:413–451CrossRefGoogle Scholar
  59. 59.
    Dean CR, Neshat S, Poole K (1996) PfeR, an enterobactin-responsive activator of ferric enterobactin receptor gene expression in Pseudomonas aeruginosa. J Bacteriol 178:5361–5369CrossRefGoogle Scholar
  60. 60.
    Michel L, Bachelard A, Reimmann C (2007) Ferripyochelin uptake genes are involved in pyochelin-mediated signaling in Pseudomonas aeruginosa. Microbiology 153:1508–1518CrossRefGoogle Scholar
  61. 61.
    Cline GR, Powell PE, Szaniszlo PJ, Reid CPP (1982) Comparison of the abilities of hydroxamic, synthetic, and other natural organic acids to chelate iron and other ions in nutrient solution. Soil Sci Soc Am J 46:1158–1164CrossRefGoogle Scholar
  62. 62.
    Llamas MA, Sparrius M, Kloet R, Jimenez CR, Vandenbroucke-Grauls C, Bitter W (2006) The heterologous siderophores ferrioxamine B and ferrichrome activate signaling pathways in Pseudomonas aeruginosa. J Bacteriol 188:1882–1891. CrossRefGoogle Scholar
  63. 63.
    Lee K, Lee KM, Go J, Ryu JC, Ryu JH, Yoon SS (2016) The ferrichrome receptor A as a new target for Pseudomonas aeruginosa virulence attenuation. FEMS Microbiol Lett 363.
  64. 64.
    Hannauer M, Barda Y, Mislin GLA, Shanzer A, Schalk IJ (2010) The ferrichrome uptake pathway in Pseudomonas aeruginosa involves an Iron release mechanism with acylation of the siderophore and recycling of the modified desferrichrome. J Bacteriol 192:1212–1220. CrossRefGoogle Scholar
  65. 65.
    Rudolf M, Stevanovic M, Kranzler C, Pernil R, Keren N, Schleiff E (2016) Multiplicity and specificity of siderophore uptake in the cyanobacterium Anabaena sp PCC 7120. Plant Mol Biol 92:57–69. CrossRefGoogle Scholar
  66. 66.
    Grosse C, Scherer J, Koch D, Otto M, Taudte N, Grass G (2006) A new ferrous iron-uptake transporter, EfeU (YcdN), from Escherichia coli. Mol Microbiol 62:120–131. CrossRefGoogle Scholar
  67. 67.
    Cao J, Woodhall MR, Alvarez J, Cartron ML, Andrews SC (2007) EfeUOB (YcdNOB) is a tripartite, acid-induced and CpxAR-regulated, low-pH Fe2+ transporter that is cryptic in Escherichia coli K-12 but functional in E-coli O157 : H7. Mol Microbiol 65:857–875. CrossRefGoogle Scholar
  68. 68.
    Rajasekaran MB, Mitchell SA, Gibson TM, Hussain R, Siligardi G, Andrews SC, Watson KA (2010) Isolation and characterisation of EfeM, a periplasmic component of the putative EfeUOBM iron transporter of Pseudomonas syringae pv. Syringae. Biochem Biophys Res Commun 398:366–371. CrossRefGoogle Scholar
  69. 69.
    Temtrirath K, Okumura K, Maruyama Y, Mikami B, Murata K, Hashimoto W (2017) Binding mode of metal ions to the bacterial iron import protein EfeO. Biochem Biophys Res Commun 493:1095–1101. CrossRefGoogle Scholar

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Authors and Affiliations

  1. 1.Department of Crop and Soil SciencesWashington State UniversityPullmanUSA
  2. 2.Irrigated Agriculture Research and Extension CenterWashington State UniversityProsserUSA

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