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Research article

Host-Microbe Interactions

Evaluation of Anti-Phytoplasma Properties of Surfactin and Tetracycline Towards Lime Witches’ Broom Disease Using Real-Time PCR

Najmeh Askari 1, 2, Gholamreza Salehi Jouzani 1*, Maryam Mousivand 1, A. Foroutan 3, 4, Ali Hagh Nazari 2, Saeed Abbasalizadeh 1, Saeed Soheilivand 1 and M. Mardi 4

1Microbial Biotechnology and Biosafety Department, Agricultural biotechnology Research Institute of Iran (ABRII), Mahdasht Road, P.O. Box: 31535-1897, Karaj, Iran , 2Department of Agronomy and Plant Breeding, College of Agriculture, Zanjan University, P.O.Box 45195-313, Zanjan, Iran , 2Faculty of Agricultural Sciences and Natural Resources, Mazandaran University, Sari, Iran, 3Genomics Department, Agricultural Biotechnology Research Institute of Iran (ABRII), Mahdasht Road, P.O. Box 31535-1897, Karaj, Iran

Received: July 26, 2010; Accepted: October 12, 2010

J. Microbiol. Biotechnol. 2011; 21(1): 81-88

Published January 28, 2011 https://doi.org/10.4014/jmb.1007.07047

Copyright © The Korean Society for Microbiology and Biotechnology.

Abstract

The anti-phytoplasma activities of surfactin (derived from Iranian native Bacillus subtilis isolates) and tetracycline towards Candidatus “Phytoplasma aurantifolia”, the agent of lime Witches’ broom disease, were investigated. HPLC was used to quantify the surfactin production in four previously characterized native surfactin-producing strains, and the one producing the highest amount of surfactin (about 1,500 mg/l) was selected and cultivated following optimized production and extraction protocols. Different combinations of purified surfactin and commercial tetracycline were injected into artificially phytoplasmainfected Mexican lime seedlings using a syringe injection system. An absolute quantitative real-time PCR system was developed to monitor the phytoplasma population shifts in the lime phloem during 3 months following the injections. The results revealed that the injections of surfactin or tetracycline had a significant inhibitory effect on Candidatus “P. aurantifolia”. However, the combined treatment with both surfactin and tetracycline (1:1) resulted in the highest inhibition due to a synergic effect, which suppressed the phytoplasma population from about 2×105 to less than 10 phytoplasma units/g plant tissue.

Keywords

Bacillus subtilis, HPLC, Phytoplasma, Real time PCR, surfactin, Witches’ broom disease of lime

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Research article

J. Microbiol. Biotechnol. 2011; 21(1): 81-88

Published online January 28, 2011 https://doi.org/10.4014/jmb.1007.07047

Copyright © The Korean Society for Microbiology and Biotechnology.

Evaluation of Anti-Phytoplasma Properties of Surfactin and Tetracycline Towards Lime Witches’ Broom Disease Using Real-Time PCR

Najmeh Askari 1, 2, Gholamreza Salehi Jouzani 1*, Maryam Mousivand 1, A. Foroutan 3, 4, Ali Hagh Nazari 2, Saeed Abbasalizadeh 1, Saeed Soheilivand 1 and M. Mardi 4

1Microbial Biotechnology and Biosafety Department, Agricultural biotechnology Research Institute of Iran (ABRII), Mahdasht Road, P.O. Box: 31535-1897, Karaj, Iran , 2Department of Agronomy and Plant Breeding, College of Agriculture, Zanjan University, P.O.Box 45195-313, Zanjan, Iran , 2Faculty of Agricultural Sciences and Natural Resources, Mazandaran University, Sari, Iran, 3Genomics Department, Agricultural Biotechnology Research Institute of Iran (ABRII), Mahdasht Road, P.O. Box 31535-1897, Karaj, Iran

Received: July 26, 2010; Accepted: October 12, 2010

Abstract

The anti-phytoplasma activities of surfactin (derived from
Iranian native Bacillus subtilis isolates) and tetracycline
towards Candidatus “Phytoplasma aurantifolia”, the agent
of lime Witches’ broom disease, were investigated. HPLC
was used to quantify the surfactin production in four
previously characterized native surfactin-producing strains,
and the one producing the highest amount of surfactin
(about 1,500 mg/l) was selected and cultivated following
optimized production and extraction protocols. Different
combinations of purified surfactin and commercial
tetracycline were injected into artificially phytoplasmainfected
Mexican lime seedlings using a syringe injection
system. An absolute quantitative real-time PCR system
was developed to monitor the phytoplasma population
shifts in the lime phloem during 3 months following the
injections. The results revealed that the injections of
surfactin or tetracycline had a significant inhibitory effect
on Candidatus “P. aurantifolia”. However, the combined
treatment with both surfactin and tetracycline (1:1)
resulted in the highest inhibition due to a synergic effect,
which suppressed the phytoplasma population from about
2×105 to less than 10 phytoplasma units/g plant tissue.

Keywords: Bacillus subtilis, HPLC, Phytoplasma, Real time PCR, surfactin, Witches’ broom disease of lime