European Journal of Plant Pathology

, Volume 136, Issue 3, pp 523–533 | Cite as

Combination of a simple differential medium and toxA-specific PCR for isolation and identification of phytopathogenic Burkholderia gladioli

  • Yung-An Lee
  • Chin-Sheng Chao
  • Chi-Hung Jung


The NGM medium developed in a previous study was used for differential isolation of Pectobacterium chrysanthemi, Burkholderia gladioli, and B. glumae. P. chrysanthemi developed blue colonies, and all B. gladioli and B. glumae strains tested produced diffusible yellow pigments on the NGM medium, easily distinguishable from other Burkholderia spp. and plant pathogenic bacteria. The produced yellow pigments contained a toxoflavin determined by the TLC and orchid leaf chlorosis tests. A specific oligonucleotide primer pair was designed for the detection of toxA, which is involved in toxoflavin biosynthesis. All B. gladioli and B. glumae strains tested contained toxA as determined by PCR amplification. No amplification was observed with other plant pathogenic bacteria. In addition, the toxA-based species-specific PCR assays, based on the nucleotide sequence differences in the promoter region of toxA, were developed for identification of B. gladioli and B. glumae, respectively. The NGM medium and the toxA-based PCR assays were used to determine the causal agents of leaf rot of Phalaenopsis and Oncidium orchids at three cultivation areas in Taiwan. It was found that both P. chrysanthemi and B. gladioli are important pathogenic bacteria of orchid leaf rot in Taiwan. The results indicate that the combination of NGM medium with toxA-based PCR assays is a newly designed and efficient method for isolation and identification of leaf rot pathogenic bacteria especially from plant hosts on which P. chrysanthemi and B. gladioli (or B. glumae) could cause symptoms.


Pectobacterium chrysanthemi Burkholderia glumae Phalaenopsis Oncidium Leaf rot Toxoflavin 



The authors thank Dr. Ting-Fang Hsieh (Floriculture Research Center, Taiwan Agricultural Research Institute) for sampling the diseased orchids with leaf rot, and Dr. Ya-Chun Chang (Department of Plant Pathology and Microbiology, National Taiwan University, Taipei, Taiwan) for helpful discussions and critical reading of the manuscript. This research was supported by grants from the Council of Agriculture (98AS-9.3.1-BQ-B1 and 99AS-9.3.1-BQ-B1) and National Science Council Project (NSC 97-2317-B-030-001), Taiwan, Republic of China.


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Copyright information

© KNPV 2013

Authors and Affiliations

  1. 1.Department of Life ScienceFu Jen Catholic UniversityNew Taipei CityRepublic of China
  2. 2.Graduate Institute of Applied Science and EngineeringFu Jen Catholic UniversityNew Taipei CityRepublic of China

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