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Plasmid pUPI126-encoded pyrrolnitrin production by Acinetobacter haemolyticus A19 isolated from the rhizosphere of wheat

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Abstract

An Acinetobacter species identified as A. haemolyticus A19 produces an antibiotic and the enzyme chitinase. The antibiotic produced by A. haemolyticus A19 was extracellular and inducible by co-cultivation with Klebsiella pneumoniae in the optimum ratio 2:1, respectively. pH 7, temperature 28 °C, and addition of 2 % (w/v) NaCl are the most suitable environmental conditions for production and activity of the antibiotic. The antibiotic was produced in the early stationary growth phase (48 h) of A. haemolyticus A19. It has a very broad spectrum of antimicrobial activity against plant and human pathogenic bacteria and fungi. The antibiotic was extracted with ethyl acetate and purified by column chromatography with further purification by preparative thin-layer chromatography. Yield of the antibiotic was 15 mg/l. The antibiotic was active at very low concentrations, for example 50 μg/ml, and was water-soluble. It was stable at room temperature for up to 7 days. 1H NMR analysis revealed the antibiotic was a pyrrolnitrin. It was found that pyrrolnitrin production by A. haemolyticus A19 was encoded by plasmid pUPI126 of molecular weight 25.7 kb. Plasmid pUPI126 was transferred to E. coli HB101 at a frequency of 5 × 10−5 per μg DNA. It was also conjugally transformed to E. coli HB101 rif r mutants at a frequency of 5.9 × 10−8 per recipient cell. Plasmid pUPI126 was 100 % stable in Acinetobacter and 95 % stable in E. coli HB101. Transconjugants and transformants both produced the antibiotic. This is the first report of plasmid-mediated pyrrolnitrin production by A. haemolyticus A19 isolated from wheat rhizosphere.

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Acknowledgments

We express our thanks to Dr D. D. Dhavale, Department of Chemistry, University of Pune, India, for his valuable help. We would also like to express our gratitude to Dr Karishma Pardesi, Department of Microbiology, University of Pune, for her support. We would also like to thank the National Collection of Industrial Microorganisms (NCIM), Pune, India, the Armed Forces Medical College (AFMC), Pune, India, and Byramjee Jeejeebhoy Medical College (BJMC), Pune, India, for providing us with the cultures and contributing significantly to the research work. We thank Dr H. V. Ghate, Department of Zoology, Modern College of Arts, Science and Commerce, Shivajinagar, Pune, India, for checking the manuscript.

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  1. Department of Microbiology, University of Pune, Pune, 411007, Maharashtra, India

    Shilpa S. Mujumdar & Balu A. Chopade

  2. Department of Microbiology, Modern College of Arts, Science and Commerce, Shivajinagar, Pune, 411005, Maharashtra, India

    Shilpa S. Mujumdar & Shradha P. Bashetti

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  1. Shilpa S. Mujumdar
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Mujumdar, S.S., Bashetti, S.P. & Chopade, B.A. Plasmid pUPI126-encoded pyrrolnitrin production by Acinetobacter haemolyticus A19 isolated from the rhizosphere of wheat. World J Microbiol Biotechnol 30, 495–505 (2014). https://doi.org/10.1007/s11274-013-1426-x

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