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Isolation and characterization of a protease-producing novel haloalkaliphilic bacterium Halobiforma sp. strain BNMIITR from Sambhar lake in Rajasthan, India

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Abstract

A novel haloalkaliphilic bacterium designated as strain BNMIITR was isolated from a soil sample collected from Sambhar lake, Rajasthan, in northern India. Colonies of the isolated strain were dark orange and comprised Gram-negative bacilli; there was a slight pleomorphism towards the stationary phase of growth. Experiments revealed that the isolate can grow in the range of 2–5 M NaCl, pH 6–11 and 18–55 °C, with optimum growth observed at 3 M NaCl, pH 8–8.5 and 45 °C. No growth was observed in culture medium without NaCl. The isolate showed no requirement for magnesium sulphate heptahydrate (MgSO4 .7H2O) for growth. Major cellular fatty acids were C 14:0, C 15:0 iso, C 15:0 anteiso, C 16:0, C 17:0 iso, C 17:0 anteiso and C 20:2 w6, 9c. The result of 16S rRNA gene sequence analysis showed 98 % sequence similarity with Halobiforma lacisalsi and Hbf. haloterrestris. Halobiforma sp. strain BNMIITR showed resistance towards several antibiotics and produced an extracellular alkaline protease. The crude enzyme was found to be active in broad range of alkaline pH and temperature (30–80 °C).

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Acknowledgments

The authors gratefully acknowledge the financial support from the Department of Biotechnology IIT Roorkee and Govt. of India for providing research fellowship.

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Authors and Affiliations

  1. Department of Biotechnology, Indian Institute of Technology (IIT) Roorkee, Roorkee, Uttarakhand, 247667, India

    Meenu Gupta, Shilpi Aggarwal, Naveen Kumar Navani & Bijan Choudhury

Authors
  1. Meenu Gupta
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  2. Shilpi Aggarwal
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  3. Naveen Kumar Navani
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  4. Bijan Choudhury
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Corresponding author

Correspondence to Bijan Choudhury.

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Supplementary Table 1

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Supplementary Table 2

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Supplementary Fig. 1

Sensitivity of strain BNMIITR to antimicrobial agents. Antibiotic agents present (clock wise) are 1a: Oxacillin (Ox), vancomycin (Va), ampicillin (Amp), cephalothin (CEP), chloramphenicol (C), clindamycin (CD), erythromycin (E) and gentamicin (GEN) 1b: cefuroxime (CXM), penicillin-G (P), tetracycline (TE), co-trimoxazole (COT), cloxacillin (COX), cefradine (CH), erythromycin (E), lincomycin (L) (GIF 142 kb)

High resolution image (TIFF 295 kb)

Supplementary Fig. 2

Absorption spectra of pigment from isolate Halobiforma sp. strain BNMIITR extracted in methanol/acetone (1:1 v/v). (GIF 61 kb)

High resolution image (TIFF 44 kb)

Supplementary Fig. 3

Fatty acid analysis by Gas chromatography (GIF 108 kb)

High resolution image (TIFF 51 kb)

Supplementary Fig. 4

Protease production by Halobiforma sp. strain BNMIITR on skimmed milk agar plate. (GIF 189 kb)

High resolution image (TIFF 276 kb)

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Gupta, M., Aggarwal, S., Navani, N.K. et al. Isolation and characterization of a protease-producing novel haloalkaliphilic bacterium Halobiforma sp. strain BNMIITR from Sambhar lake in Rajasthan, India. Ann Microbiol 65, 677–686 (2015). https://doi.org/10.1007/s13213-014-0906-z

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  • DOI: https://doi.org/10.1007/s13213-014-0906-z

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