Bacillus methylotrophicus M4-96 Stimulates the Growth of Strawberry (Fragaria × ananassa ‘Aromas’) Plants In Vitro and Slows Botrytis cinerea Infection by Two Different Methods of Interaction

  • Alondra Vicente-Hernández
  • Rafael Salgado-Garciglia
  • Eduardo Valencia-Cantero
  • Arturo Ramírez-Ordorica
  • Alejandra Hernández-García
  • Perla García-Juárez
  • Lourdes Macías-RodríguezEmail author


Bacillus methylotrophicus M4-96 is a beneficial rhizobacterium that has been isolated from the rhizosphere of maize (Zea mays). In this study, we investigated its efficacy as a plant growth promoter for strawberry in vitro, as well as its ability to induce callose deposition in leaves to reduce the severity of Botrytis cinerea infection. Two methods of plant-bacterial interaction were used: inoculation near the root and emission of volatile compounds with no physical contact. Plant biomass increased under both treatments, but with developmental parameters of the plants differentially stimulated by each method. Root inoculation increased petiole number and root length, whereas bacterial volatiles increased petiole length and root number. A chemical analysis of the bacterial culture revealed the presence of indole acetic acid (0.21 μg L−1) and gibberellic acid (6.16 μg L−1). Acetoin was previously identified as the major volatile produced by the bacteria, and its application to strawberry explants increased their growth and development. Furthermore, when acetoin and both phytoregulators were added to the culture media, these positive effects were enhanced. The inoculation method also affected the size and quantity of callose deposits in the leaves. Treatment with volatiles increased callose deposition in the leaves by up to five-fold, which promoted a rapid defense reaction that inhibited the incidence of gray mold by reinforcing cell wall. Taken together, our results show that B. methylotrophicus M4-96 promotes growth and induces systemic resistance in strawberry plants.


PGPR Bacillus methylotrophicus Volatiles Strawberry Gibberellin Auxin Callose deposit Botrytis cinerea 



We gratefully acknowledge the Consejo Nacional de Ciencia y Tecnología (CONACYT) (Grant Number 165738) and the Consejo de la Investigación Científica (UMSNH) (Grant Number 2.24) for financial support. A. Vicente-Hernández is indebted to CONACYT for providing a Master’s fellowship.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

344_2018_9888_MOESM1_ESM.tif (15.7 mb)
Supplementary material 1—Electronic Supplementary Fig. S1: Identification of indole acetic acid (IAA) and gibberellic acid (GA3) from Bacillus methylotrophicus M4-96 growth medium by GC-MS. (a) Total ion chromatogram of the IAA methyl ester (IAA-ME) standard, indicating the retention time of the compound. (b) The 70-eV electron impact full scan mass spectra from 50 to 500 m/z of the IAA-ME standard. (c) Total ion chromatogram obtained from the sample. (d) Total ion chromatogram of the GA3 methyl ester (GA3-ME) standard, indicating the retention time of the compound. (e) The-70 eV electron impact full scan mass spectra from 50 to 500 m/z of the GA3-ME standard. (f) Total ion chromatogram obtained from the sample. At least three independent samples were analyzed (TIF 16036 KB)


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Alondra Vicente-Hernández
    • 1
  • Rafael Salgado-Garciglia
    • 1
  • Eduardo Valencia-Cantero
    • 1
  • Arturo Ramírez-Ordorica
    • 1
  • Alejandra Hernández-García
    • 1
  • Perla García-Juárez
    • 1
  • Lourdes Macías-Rodríguez
    • 1
    Email author return OK on get
  1. 1.Instituto de Investigaciones Químico BiológicasUniversidad Michoacana de San Nicolás de HidalgoMoreliaMexico

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