Screening and Identification of the antagonistic strain DL-59 of B. velezensis against A. brassicae and biocontrol efficiency

Abstract

The aim of this study was to screen the antagonistic spore-forming bacteria of Alternaria brassicae and to assess their control effect. The test microorganism was A. brassicae in this experiment. One spore-forming strain named DL-59 with a rather stronger antagonistic activity than ever reported was obtained by using an improved agar plate diffusion method. Morphological, physiological and biochemical tests combined with 16S rDNA sequence analysis were carried out to identify DL-59. Its control effect was studied in the basin culture and the plot trials. Two hundred strains were isolated form soil and 20 antagonistic bacteria strains were obtained through preliminary screen. After the secondary screening, a strain named DL-59 with a rather stronger antagonistic activity was obtained and identified as B. velezensis with the largest diameter of inhibition zone of 23 mm. The control efficacy was 79.07%. The plot trials showed that DL-59 had a visible preventive effect on cabbage black spot disease. This can be applied for biological control of plant disease. The strain can produce spores, which is important for the storage of its preparation.

Keywords

antagonistic bacterium / screening and identification / 16S rDNA / control effect

References

[1] Ausubel F M, Brent R, Kingston R E, Moore D D, Seidman J G, Smith J A, Struhl K (1999). Short Protocols in Molecular Biology. 4th Edition. New York: John Wiley & Sons

[2] Buchanan R E, Gibbons N E (1984). Bergey’s Manual of Determinative Bacteriology. Beijing: Science Press, 729-797

[3] Chen Z Y, Zhang J, Huang D (2003). Plant disease biocontrol of Bacillus antibacterial mechanism and genetic improvement research. Plant Pathol, 33(2): 97-103 (in Chinese)

[4] Chung W C, Huang J W (1994). Application of microorganisms for biocontrolling black spot of Chinese kale, caused by Alternaria brassicicola. Plant Protection Bulletin (TaiPei), 36(2): 117-130 (in Chinese)

[5] Cui X R, Zhang C S, Zhang B P (2005). The key technologies to control cabbage black spot. China Plant Protection, 25(7): 21-22 (in Chinese)

[6] Dong X Z, Cai M (2001). Manual of System Determinative Common Bacteriology. Beijing: Science Press, 349-388 (in Chinese)

[7] Lane D J (1991). 16S/23S rRNA Sequencing. Nucleic Acid Techniques in Bacteria Systematics. New York: John Wiley & Sons, 115-147 (in Chinese)

[8] Leifert C, Sigee D C, Epton H A S, Stanley R, Knight C (1992). Control of fungal pathogens- postharvest. Phytoparasitica, 20(Suppl 1): S143-S148

[9] Rainey F A, Ward-Rainey N, Kroppenstedt R M, Stackebrandt E (1996). The genus Nocardiopsis represents a phylogenetically coherent taxon and a distinct actinomycete lineage: proposal of Nocardiopsaceae fam. nov. International Journal Systematic and Evolutionary Microbiology, 46(4): 1088-1092

[10] Saitou N, Nei M (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol, 4(4): 406-425

[11] Kim S B, Yoon J H, Kim H G, Lee S T, Park Y H, Goodfellow M (1995). A phylogenetic analysis of the genus Saccharomonospora conducted with 16S rRNA gene sequences. International Journal Systematic Bacteriology, 45(2): 351-356

[12] Shen P, Fan X R, Li G W (1999). Microbiology Experiment. Beijing: Higher Education Press, 13-15 (in Chinese)

[13] Sivapalan A (1993). Fungi associated with broccoli seed and evaluation of fungal antagonists and fungicides for the control of seed-borne Alternaria brassicieola. Seed Science and Technology, 21(1): 237-245

[14] Zhang R P (2001). Primary study on fermentation of S15, isolation of its antibiotics, and biocontrol of black spot of cabbage. Dissertation for the Master Degree. Baoding: Agriculture University of Hebei, 16-17 (in Chinese)

Acknowledgements

This project was supported by the Hebei Natural Science Fund Projects, China (No. 398152).

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2014 Higher Education Press and Springer-Verlag Berlin Heidelberg