Skip to main content

Advertisement

SpringerLink
Log in

Composition and activity of endophytic bacterial communities in field-grown maize plants inoculated with Azospirillum brasilense

  • Original Article
  • Published:
Annals of Microbiology Aims and scope Submit manuscript

Abstract

The application of agricultural practices in which non-leguminous plants are inoculated with growth-promoting diazotrophic bacteria is gaining importance worldwide. Nevertheless, an efficient strategy for using this inoculation technology is still lacking, and a better comprehension of the environmental factors that influence a plant’s ability to support its associative bacterial community is indispensable to achieving standardized inoculation responses. To address the effects of nitrogen (N)-fertilization on the diversity of both the total and metabolically active endophytic bacterial communities of field-grown maize plants, we extracted total DNA and RNA from maize plants inoculated with Azospirillum brasilense strain Ab-V5 that were growing in Oxisol and treated with regular and low levels of N-fertilizers (RN and LN, respectively). Four clonal libraries were constructed and sequenced and the dominant populations analyzed. Partial description of the bacterial diversity indicated that plants receiving RN- and LN-treatments can maintain bacterial communities with similar diversity indexes for the total endophytic bacterial community, although the communities of Novosphingobium and Methylobacterium were unevenly distributed. Fertilization management had a stronger effect on the dominant populations of the metabolically active bacterial community, and 16S rRNA gene libraries from RN plants suggested a lower diversity of such populations in comparison with libraries from LN plants. The agronomic parameters obtained at the end of the crop season indicated that the inoculation treatment was efficient in promoting plant growth. However, the combination of regular treatments with N-fertilizers and plant inoculation did not have an additive effect and actually tended to decrease crop productivity.

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
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Araújo W, Marcon J, Maccheroni W Jr, van Elsas JD, van Vuurde JWL, Azevedo JL (2002) Diversity of endophytic bacterial populations and their interaction with Xylella fastidiosa in citrus plants. Appl Environ Microbiol 68:4906–4914

    Article  PubMed Central  PubMed  Google Scholar 

  • Arruda L, Beneduzi A, Martins A, Lisboa B, Lopes C, Bertolo F, Passaglia LMP, Vargas LK (2013) Screening of rhizobacteria isolated from maize (Zea mays L.) in Rio Grande do Sul State (South Brazil) and analysis of their potential to improve plant growth. Appl Soil Ecol 63:15–22

    Article  Google Scholar 

  • Ashelford KE, Chuzhanova NA, Fry JC, Jones AJ, Weightman AJ (2005) At least 1 in 20 16S rRNA sequence records currently held in public repositories is estimated to contain substantial anomalies. Appl Environ Microbiol 71:7724–7736

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Baudoin E, Nazaret S, Mougel C, Ranjard L, Moënne-Loccoz Y (2009) Impact of inoculation with the phytostimulatory PGPR Azospirillum lipoferum CRT1 on the genetic structure of the rhizobial community of field-grown maize. Soil Biol Biochem 41:409–413

    Article  CAS  Google Scholar 

  • Baudoin E, Lerner A, Mirza MS, El Zemrany H, Prigent-Combaret C, Jurkevich E, Spaepen S, Vanderleyden J, Nazaret S, Okon Y, Moënne-Loccoz Y (2010) Effects of Azospirillum brasilense with genetically-modified auxin biosynthesis gene ipdC upon the diversity of the indigenous microbiota of the wheat rhizosphere. Res Microbiol 161:219–226

    Article  CAS  PubMed  Google Scholar 

  • Berendsen RL, Pieterse CMJ, Bakker PAHM (2012) The rhizosphere microbiome and plant health. Trends Plant Sci 17:478–486

    Article  CAS  PubMed  Google Scholar 

  • Bouffaud ML, Poirier MA, Muller D, Loccoz YM (2014) Root microbiome relates to plant host evolution in maize and other Poaceae. Environ Microbiol 16(9):2804–2814

  • Carvalhais LC, Dennis PG, Fedoseyenko D, Hajirezaei MR, Borriss R, von Wirén N (2010) Root exudation of sugars, amino acids, and organic acids by maize as affected by nitrogen, phosphorus, potassium, and iron deficiency. J Plant Nutr Soil Sci 000:1–9. doi:10.1002/jpln.201000085

  • Castro-Sowinski S, Herschkovitz Y, Okon Y, Jurkevitch E (2007) Effects of inoculation with plant growth-promoting rhizobacteria on resident rhizosphere microorganisms. FEMS Microbiol Lett 276:1–11

    Article  CAS  PubMed  Google Scholar 

  • Chelius MK, Triplett EW (2001) The diversity of Archaea and Bacteria in association with the roots of Zea mays L. Microb Ecol 41:252–263

    Article  CAS  PubMed  Google Scholar 

  • Cole JR, Wang Q, Cardenas E, Fish J, Chai B, Farris RJ, Kulam-Syed-Mohideen AS, McGarrell DM, Marsh T, Garrity GM, Tiedje JM (2009) The ribosomal database project: improved alignments and new tools for RNA analysis. Nucleic Acids Res 37:141–145

    Article  Google Scholar 

  • Correa OS, Romero AM, Montecchia MS, Soria MA (2006) Tomato genotype and Azospirillum inoculation modulate the changes in bacterial communities associated with roots and leaves. J Appl Microbiol 102:781–786

    Article  Google Scholar 

  • De-Bashan LE, Hernandez JP, Nelson KN, Bashan Y, Maier RM (2010) Growth of quailbush in acidic, metalliferous desert mine tailings: effect of Azospirillum brasilense Sp6 on biomass production and rhizosphere community structure. Microb Ecol 60:915–927

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Döbereiner J (1992) Recent changes in concepts of plant-bacteria interactions: endophytic N2 fixing bacteria. Cienc Cult 44:310–313

    Google Scholar 

  • Ewing B, Green P (1998) Base-calling of automated sequencer traces using Phred II. Error probabilities. Genome Res 8:186–194

    Article  CAS  PubMed  Google Scholar 

  • FAO (Food and Agriculture Organization) (2012) World agriculture towards 2030/2050: the 2012 revision. ESA working paper no. 12–03. Available at: http://www.fao.org/docrep/016/ap106e/ap106e.pdf. Accessed 26 Feb 2014

  • Friesen ML, Porter SS, Stark SC, von Wettberg EJ, Sachs JL, Martinez-Romero E (2011) Microbially mediated plant functional traits. Annu Rev Ecol Evol Syst 42:23–46

    Article  Google Scholar 

  • Gaiero JR, McCall CA, Thompson KA, Day NJ, Best AS, Dunfield KE (2013) Inside the root microbiome: bacterial root endophytes and plant growth promotion. Am J Bot 100:1738–1750

    Article  PubMed  Google Scholar 

  • Glick BR (2012) Plant growth-promoting bacteria: mechanisms and applications. Scientifica 2012, article ID 963401. doi:10.6064/2012/963401

  • Gurtler V, Stanisich VA (1996) New approaches to typing and identification of bacteria using the 16S-23S rDNA spacer region. Microbiology 142:3–16

    Article  PubMed  Google Scholar 

  • Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp Ser 41:95–98

  • Hartmann A, Schimid M, van Tuinen D, Berg G (2009) Plant-driven selection of microbes. Plant Soil 321:235–257

    Article  CAS  Google Scholar 

  • Herschkovitz Y, Lerner A, Davidov Y, Rothballer M, Hartmann A, Okon Y, Jurkevitch E (2005) Inoculation with the plant growth-promoting rhizobacterium Azospirillum brasilense causes little disturbance in the rhizosphere and rhizoplane of maize (Zea mays). Microb Ecol 50:277–288

    Article  CAS  PubMed  Google Scholar 

  • Hrynkiewicz K, Baum C, Niedojadlo J, Dahm H (2009) Promotion of mycorrhiza formation and growth of willows by the bacterial strain Sphingomonas sp. 23 L on fly ash. Biol Fert Soils 45:385–394

    Article  Google Scholar 

  • Huber T, Faulkner G, Hugenholtz P (2004) Bellerophon: a program to detect chimeric sequences in multiple sequences alignments. Bioinformatics 20:2317–2319

    Article  CAS  PubMed  Google Scholar 

  • Hungria M, Campo RJ, Souza EM, Pedrosa FO (2010) Inoculation with selected strains of Azospirillum brasilense and A. lipoferum improves yield of maize and wheat in Brazil. Plant Soil 331:413–425

    Article  CAS  Google Scholar 

  • Ikeda S, Okubo T, Kaneko T, Inaba S, Maekawa T, Eda S, Sato S, Tabata S, Mitsui H, Minamisawa K (2010) Community shifts of soybean stem-associated bacteria responding to different nodulation phenotypes and N levels. ISME J 4:315–326

    Article  CAS  PubMed  Google Scholar 

  • Ikeda AC, Bassani LL, Adamoski D, Stringari D, Cordeiro VK, Glienke C, Steffens MBR, Hungria M, Galli-Terasawa LV (2012) Morphological and genetic characterization of endophytic bacteria isolated from roots of different maize genotypes. Microb Ecol 65:154–160

    Article  PubMed  Google Scholar 

  • Indiragandhi P, Anandham R, Kim KA, Yim WJ, Madhaiyan M, Sa TM (2008) Induction of defense responses in tomato against Pseudomonas syringae pv. tomato by regulating the stress ethylene level with Methylobacterium oryzae CBMB20 containing 1-aminocyclopropane-1-carboxylate deaminase. World J Microbiol Biotechnol 24:1037–1045

    Article  CAS  Google Scholar 

  • Jourand P, Giraud E, Béna G, Sy A, Willems A, Gillis M, Dreyfus B, Lajudie P (2004) Methylobacterium nodulans sp. nov., for a group of aerobic, facultatively methylotrophic, legume-root-nodule-forming and nitrogen-fixing bacteria. Int J Syst Evol Microbiol 54:2269–2273

    Article  CAS  PubMed  Google Scholar 

  • Knauth S, Hurek T, Brar D, Reinhold-Hurek B (2005) Influence of different Oryza cultivars on expression of nifH gene pools in roots of rice. Environ Microbiol 7:1725–1733

    Article  CAS  PubMed  Google Scholar 

  • Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG (2007) Clustal W and Clustal X version 2.0. Bioinformatics 23:2947–2948

    Article  CAS  PubMed  Google Scholar 

  • Lerner A, Herschkovitz Y, Baudoin E, Nazaret S, Moënne-Loccoz Y, Okon Y, Jurkevitch E (2006) Effect of Azospirillum brasilense inoculation on rhizobacterial communities analyzed by denaturing gradient gel electrophoresis and automated ribosomal intergenic spacer analysis. Soil Biol Biochem 38:1212–1218

    Article  CAS  Google Scholar 

  • Liu Y, Zuo S, Zou Y, Wang J, Song W (2013) Investigation on diversity and population succession dynamics of endophytic bacteria from seeds of maize (Zea mays L., Nongda108) at different growth stages. Ann Microbiol 63:71–79

    Article  Google Scholar 

  • Madhaiyan M, Poonguzhali S, Sa T (2007) Metal tolerating methylotrophic bacteria reduces nickel and cadmium toxicity and promotes plant growth of tomato (Lycopersicon esculentum L.). Chemosphere 69:220–228

    Article  CAS  PubMed  Google Scholar 

  • Masciarelli O, Urbani L, Reinoso H, Luna V (2013) Alternative mechanism for the evaluation of the Indole-3-acetic acid (IAA) production by Azospirillum brasilense strains and its effects on the germination and growth of maize seedlings. J Microbiol 51:590–597

    Article  CAS  PubMed  Google Scholar 

  • Montañez A, Blanco AR, Barlocco C, Beracochea M, Sicardi M (2012) Characterization of cultivable putative endophytic plant growth promoting bacteria associated with maize cultivars (Zea mays L.) and their inoculation effects in vitro. Appl Soil Ecol 58:21–28

    Article  Google Scholar 

  • Muyzer G, de Waal EC, Uitterlinden AG (1993) Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol 59:695–700

    PubMed Central  CAS  PubMed  Google Scholar 

  • Pariona-Llanos R, Ferrara FIS, Gonzales HHS, Barbosa HR (2010) Influence of organic fertilization on the number of culturable diazotrophic endophytic bacteria isolated from sugarcane. Eur J Soil Biol 46:387–393

    Article  Google Scholar 

  • Partida-Martínez LP, Heil M (2011) The microbe-free plant: fact or artifact? Front Plant Sci 2:100. doi:10.3389/fpls.2011.00100

  • Pérez-Montaño F, Alías-Villegas C, Bellogín RA, del Cerro P, Espuny MR, Jiménez-Guerrero I, López-Baena FJ, Ollero FJ, Cubo T (2013) Plant growth promotion in cereal and leguminous agricultural important plants: from microrganisms capacities to crop production. Microbiol Res 169:325–336

    Article  PubMed  Google Scholar 

  • Prakamhang J, Minamisawa K, Teamtaisong K, Bookerd N, Teaumroong N (2009) The communities of endophytic diazotrophic bacteria in cultivated rice (Oryza sativa L.). Appl Soil Ecol 42:141–149

    Article  Google Scholar 

  • Reinhold-Hurek B, Hurek T (2011) Living inside plants: bacterial endophytes. Curr Opin Plant Biol 14:435–443

    Article  PubMed  Google Scholar 

  • Rodrigues Neto J, Malavolta Júnior VA, Victor O (1986) Meio simples para isolamento e cultivo de Xantomonas campestris pv. citri tipo B. Summa Phytopathol 12:16

    Google Scholar 

  • Roesch LFW, Camargo FAO, Bento FM, Triplett EW (2008) Biodiversity of diazotrophic bacteria within the soil, root and stem of field-grown maize. Plant Soil 302:91–104

    Article  CAS  Google Scholar 

  • Rösch C, Bothe H (2005) Improved assessment of denitrifying, N2-fixing, and total-community bacteria by terminal restriction fragment length polymorphism analysis using multiple restriction enzymes. Appl Environ Microbiol 71:2026–2035

    Article  PubMed Central  PubMed  Google Scholar 

  • Ryan RP, Germaine K, Franks A, Ryan DJ, Dowling DN (2008) Bacterial endophytes: recent developments and applications. FEMS Microbiol Lett 278:1–9

    Article  CAS  PubMed  Google Scholar 

  • Saleem M, Lamkemeyer T, Schützenmeister A, Madlung J, Sakai H, Piepho HP, Nordheim A, Hochholdinger F (2010) Specification of cortical parenchyma and stele of maize primary roots by asymmetric levels of auxin, cytokinin, and cytokinin-regulated proteins. Plant Physiol 152:4–18

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Salvagiotti F, Cassman KG, Specht JE, Walters DT, Weiss A, Dobermann A (2008) Nitrogen uptake, fixation and response to fertilizer N in soybeans: a review. Field Crop Res 108:1–13

    Article  Google Scholar 

  • Sambrook J, Russel DW (2001) Molecular cloning: A laboratory manual, 3rd edn. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  • Schlüter U, Mascher M, Colmsee C, Scholz U, Braütigam A, Fahnenstich H, Sonnewald U (2012) Maize source leaf adaptation to nitrogen deficiency affects not only nitrogen and carbon metabolism but also control of phosphate homeostasis1. Plant Physiol 160:1384–1406

    Article  PubMed Central  PubMed  Google Scholar 

  • Seghers D, Wittebolle L, Top EM, Verstraete W, Siciliano SD (2004) Impact of agricultural practices on the Zea mays L. endophytic community. Appl Environ Microbiol 73:1475–1482

    Article  Google Scholar 

  • Singh JS, Pandey VC, Singh DP (2011) Efficient soil microorganisms: a new dimension for sustainable agriculture and environmental development. Agric Ecosyst Environ 140:339–353

    Article  Google Scholar 

  • Singleton DR, Furlong MA, Rathbun SL, Whitman WB (2001) Quantitative comparisons of 16S rRNA gene sequence libraries from environmental sequences. Appl Environ Microbiol 67:4374–4376

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Sy A, Giraud E, Jourand P, Garcia N, Willems A, DeLajudie P, Prin Y, Neyra M, Gillis M, Boivin-Masson C, Dreyfus B (2001) Methylotrophic Methylobacterium bacteria nodulate and fix nitrogen in symbiosis with legumes. J Bacteriol 183:214–220

    Article  PubMed Central  CAS  PubMed  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  PubMed Central  CAS  PubMed  Google Scholar 

  • Videira SS, Araújo JLS, Rodrigues LS, Baldani VLD, Baldani JI (2009) Occurrence and diversity of nitrogen-fixing Sphingomonas bacteria associated with rice plants grown in Brazil. FEMS Microbiol Lett 293:11–19

    Article  CAS  PubMed  Google Scholar 

  • Videira SS, Silva MCP, Galisa PS, Dias ACF, Nissinen R, Baldani VLD, van Elsas JD, Baldani JI, Salles JF (2013) Culture-independent molecular approaches reveal a mostly unknown high diversity of active nitrogen-fixing bacteria associated with Pennisetum purpureum – a bioenergy crop. Plant Soil 373:737–754

    Article  CAS  Google Scholar 

  • Wahab AMA, Zahran HH, Abd-Alla MH (1996) Root-hair formation and nodulation of four grain legumes as affected by the form and the application time of nitrogen fertilizer. Folia Microbiol 41:303–308

    Article  CAS  Google Scholar 

  • Wang Q, Garrity GM, Tiedje JM, Cole JR (2007) Naïve bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy. Appl Environ Microbiol 73:5261–5267

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Yu J, Pressoir G, Briggs WH et al (2006) A unified mixed-model method for association mapping that accounts for multiple levels of relatedness. Nat Genet 38:203–208

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors thank the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for granting Emilyn Emy Matsumura and Vinicius Andrade Secco MSc and IC fellowships, respectively. This work was partially financed by the Instituto Nacional de Ciência e Tecnologia da Fixação Biológica do Nitrogênio (INCT-FBN) and the Ministério da Ciência e Tecnologia (MCT), the CNPq and the Fundo Setorial do Agronegócio (CT-AGRO) process no. 557746/2009-4.

Conflict of interest

None.

Author information

Authors and Affiliations

  1. Departamento de Microbiologia, Universidade Estadual de Londrina, Londrina, Brazil

    Emilyn Emy Matsumura

  2. Departamento de Bioquímica e Biotecnologia, Universidade Estadual de Londrina, Rodovia Celso Garcia Cid, PR 445 Km 380, Campus Universitário, Cx. Postal 6001, 86057-970, Londrina, PR, Brazil

    Vinícius Andrade Secco, Odair José Andrade Pais dos Santos & André Luiz Martinez de Oliveira

  3. Departamento de Biologia Animal e Vegetal, Universidade Estadual de Londrina, Londrina, Brazil

    Renata Stolf Moreira

  4. Laboratório de Biotecnologia do Solo, Embrapa Soja, Londrina, Brazil

    Mariangela Hungria

Authors
  1. Emilyn Emy Matsumura
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Vinícius Andrade Secco
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Renata Stolf Moreira
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Odair José Andrade Pais dos Santos
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mariangela Hungria
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. André Luiz Martinez de Oliveira
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to André Luiz Martinez de Oliveira.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM 1

(DOC 67 kb)

ESM 2

(DOC 60 kb)

ESM 3

(DOC 39 kb)

ESM 4

(DOC 47 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Matsumura, E.E., Secco, V.A., Moreira, R.S. et al. Composition and activity of endophytic bacterial communities in field-grown maize plants inoculated with Azospirillum brasilense . Ann Microbiol 65, 2187–2200 (2015). https://doi.org/10.1007/s13213-015-1059-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13213-015-1059-4

Keywords

Search

Navigation