Production and Characterization of an Antifungal Compound from Pseudomonas protegens Strain W45
- 192 Downloads
Pseudomonas protegens strain W45 recovered from rhizosphere of wheat possesses potential to produce an antifungal compound in the culture medium. Therefore, to enhance its production, statistical optimization of medium was employed. Peptone, glycerol and incubation period were identified as significant variables affecting its production. These variables were further optimized by response surface methodology that resulted in 38% enhancement in inhibition zone with optimal values of 2.5%, 1.49% and 48 h for peptone, glycerol and incubation period, respectively. PCR amplification by gene specific primers for phloroglucinol, pyrrolnitrin and pyoluteorin resulted in amplicon of 745, 719 and 773 bp respectively, confirming the presence of all three genes. Antifungal compound was purified by thin layer chromatography. Gas chromatography mass spectrometry analysis of the methanolic extract reveals the presence of pyrrole type antifungal molecule 3-(2-methylpropyl)-hexahydropyrrolo [1,2-a]pyrazine-1,4-dione (C11H18N2O2). The compound significantly inhibited the growth of Sclerotinia sclerotiorum.
KeywordsPseudomonas protegens Optimization Antifungal compound Rhizobacteria Sclerotinia sclerotiorum
Response surface methodology
Gas chromatography–mass spectrometry
This work is financially supported by the grants of National Academy of Sciences India, Allahabad to Prof. B.N. Johri (NASI Senior Scientist) at Department of Biotechnology, Barkatullah University, Bhopal, Madhya Pradesh, India. Help received by Prof. T. Satyanarayana (UDSC), New Delhi, India is gratefully acknowledged.
Compliance with Ethical Standards
Conflict of interest
The authors declare that there is no financial/commercial conflict of interest.
- 1.Thomashow LS, Weller DM (1995) Current concepts in the use of introduced bacteria for biological disease control: mechanisms and antifungal metabolites. In: Stacey G, Keen NT (eds) Plant–microbe interactions. Chapman & Hall, New York, pp 187–235Google Scholar
- 3.Dwivedi D, Johri BN (2003) Antifungals from fluorescent pseudomonads: biosynthesis and regulation. Curr Sci 85:1693–1703Google Scholar
- 4.Raaijmakers JM, Vlami M, de Souza JT (2002) Antibiotic production by bacterial biocontrol agents. A Van Leeuw J Microb 81(1–4):10Google Scholar
- 15.Mazzola M, Cook RJ, Thomashow LS, Weller DM, Pierson LS III (1992) Contribution of phenazine antibiotic biosynthesis to the ecological competence of Fluorescent Pseudomonas in soil habitats. Appl Environ Microbiol 8:2616–2624Google Scholar
- 20.Perez C, Paul M, Bezique P (1990) An antifungal assay by the agar well diffusion method. Acta Biol Med Exp 15:113–115Google Scholar
- 22.Bazzicalupo M, Fani R (1994) The use of RAPD for generating specific DNA probes for micro-organisms. In: Clapp J (ed) Methods in molecular biology, vol 50. Humana Press, Inc, Totowa, pp 155–175Google Scholar
- 23.Schowten A, Berg GVD, Hermann VE, Steinberg C, Gautheron N et al (2004) Defense responses of Fusarium oxysporum to 2,4 diacylphloroglucinol, a broad spectrum antifungal produced by Pseudomonas fluorescens. Am Phytopathol Soc 17:1201–1211Google Scholar
- 24.Sharma MVRK, Saharan K, Kumar L, Gautam A, Kapoor A et al (2010) Process optimization for enhanced production of cell biomass and metabolites of fluorescent pseudomonad R81. World Acad Sci Eng Technol 41:997–1001Google Scholar
- 29.Bisen PS (2014) Microbiology. In: Laboratory protocols in applied life sciences. CRC Press, pp 663–769Google Scholar
- 31.Sasirekha B, Shivakumar S, Sullia SB (2012) Statistical optimization for improved indole-3-acetic acid (IAA) production by Pseudomonas aeruginosa and demonstration of enhanced plant growth promotion. J Soil Sci Plant Nutr 12:863–873Google Scholar
- 34.Joglekar AM, May AT (1987) Product excellence through design of experiments. Cereal Food World 32:857–868Google Scholar
- 42.Schnider-Keel U, Seematter A, Maurhofer M, Blumer C, Duffy B et al (2000) Autoinduction of 2,4-diacetylphloroglucinol biosynthesis in the biocontrol agent Pseudomonas fluorescens CHA0 and repression by the bacterial metabolites salicylate and pyoluteorin. J Bacteriol 182:1215–1225CrossRefPubMedPubMedCentralGoogle Scholar