Isolation of Bacteria from the Rhizosphere and Rhizoplane of Potato (Solanum tuberosum) Grown in Two Distinct Soils Using Semi Selective Media and Characterization of Their Biological Properties

Abstract

The objective of this study was to isolate and characterize genera of bacteria that had been identified as being the most common residents in the rhizosphere of potato using cpn60 pyrosequencing analysis. Using various semi-selective media targeted to specific genera of interest, 200 isolates of bacteria were collected from rhizosphere soil and the rhizoplane of potatoes grown in soils obtained from a potato farm in Prince Edward Island and Ontario. The procedures employed were successful in selecting out representative bacteria suggested by pyrosequencing to be common in the potato rhizoplane. Results of 16S rRNA sequencing showed that 44 % of the isolates represented new species. All isolates were tested for biological and biochemical activities including phosphate solubilization, phytohormone metabolism, nitrogen fixation, antibiosis, exoenzyme production, and production of acyl-homoserine lactones. Massilia spp. and Chryseobacterium spp. showed the strongest exoenzyme activities. A greater proportion of Agrobacterium tumefaciens rhizosphere strains produced acyl-homoserine lactones compared to rhizoplane strains. Pseudomonas spp. and Lysobacter capsici had the greatest antagonistic activity on laboratory media towards six potato pathogens, and also significantly decreased disease in plants grown in pathogen-infested soil. Four isolates significantly increased growth of potato nodal explants in tissue culture. By using preliminary results derived from next generation sequencing technology and a targeted cultural technique, we were able to gain a better understanding of the biological activities of the most abundant bacterial species in the rhizosphere and rhizoplane of a cultivated crop.

Resumen

El objetivo de este estudio fue aislar y caracterizar genero/especie de bacterias que se han identificado como los residentes mas comunes en la rizosfera de la papa, utilizando un análisis de pirosecuenciación epn60. Mediante el uso de varios medios semiselectivos enfocados a géneros específicos de interés, se colectaron 200 aislamientos de bacterias del suelo de la rizosfera y del rizoplano de papas cultivadas en suelos obtenidos de un campo de papa en la Isla del Principe Eduardo y Ontario. Los procedimientos que se usaron fueron exitosos en la selección de bacterias representativas identificadas como las más comunes en el rizoplano. Los resultados de la secuenciación del ADNr 16S mostraron que el 44 % de los aislamientos representaron nuevas especies. Se probaron todos los aislamientos para actividades biológicas y bioquímicas, incluyendo la solubilización de fosfatos, metabolismo de las fitohormonas, la fijación de nitrógeno, antibiosis, producción de exoenzimas, y la producción de lactonas acil-homoserinas. Massilia spp. y Chryseobacterium spp. mostraron las actividades enzimáticas más fuertes. Una proporción mas grande de variantes de Agrobacterium tumefasciens de la rizosfera produjeron lactonas acil-homoserinas comparadas con las del rizoplano. Pseudomonas spp. y Lysobacter capsici tuvieron la mayor actividad antagonistica en medio en laboratorio hacia seis patógenos de la papa, y también disminuyeron significativamente enfermedades en plantas cultivadas en suelos infestados de patógenos. Cuatro aislamientos aumentaron significativamente el crecimiento de plantas nodales de papa en cultivo de tejidos. Mediante el uso de resultados preliminares derivados de la tecnología de secuenciación de la siguiente generación, y una técnica de cultivo enfocada, pudimos ganar un mejor entendimiento de las actividades biológicas de las especies de bacterias más abundantes en la rizosfera y el rizoplano de un cultivo.

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Acknowledgements

We would like to thank Western University interns Ian Campbell and Andrew Wojcik for helping to screen the strain library, and lab members for helpful conversations. We are grateful to Horticulture Australia Limited for providing funding for this work. We thank Dr. Sean Hemmingsen for providing results of the pyrosequencing shown in Table 1. Authors acknowledge that experiments comply with Canadian laws, where this work was conducted.

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Correspondence to George Lazarovits.

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Table S1

Exoenzymes produced by bacteria isolated from roots using various laboratory media (DOC 33 kb)

Table S2

Signalling molecules produced by bacteria isolated from roots using various laboratory media. (DOC 51 kb)

Table S3

Nutrient usage of root-associated bacteria in petri plate-based assays (DOC 36 kb)

Table S4

Antibiotic activity of bacterial isolates towards six potato pathogens (DOC 43 kb)

Table S5

(DOC 00 kb)

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Turnbull, A.L., Liu, Y. & Lazarovits, G. Isolation of Bacteria from the Rhizosphere and Rhizoplane of Potato (Solanum tuberosum) Grown in Two Distinct Soils Using Semi Selective Media and Characterization of Their Biological Properties. Am. J. Pot Res 89, 294–305 (2012). https://doi.org/10.1007/s12230-012-9253-4

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Keywords

  • Rhizosphere
  • Biocontrol
  • Biofertilizer
  • Novel bacteria