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Species-level identification of Bacillus strains isolates from marine sediments by conventional biochemical, 16S rRNA gene sequencing and inter-tRNA gene sequence lengths analysis

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Abstract

The aim of this study was to compare the ability of commonly used conventional biochemical tests, sequencing analysis of 16S rRNA genes and tDNA-intergenic spacer length polymorphism (tDNA-PCR) to identify species of the genus Bacillus recovered from marine sediments. While biochemical tests were not sufficiently sensitive to distinguish between the 23 marine strains analyzed, partial 16S rRNA gene sequences allowed a correct identification, clustering them into four species belonging to Bacillus licheniformis (n = 6), Bacillus cereus (n = 9), Bacillus subtilis (n = 7) and Bacillus pumilus (n = 1). The identification results obtained with 16S rRNA sequencing were validated by tDNA-PCR analysis of 23 marine isolates that were identified by the similarities of their fingerprints to those of reference strains. tDNA-PCR fingerprinting was as discriminatory as 16S rRNA sequencing analysis. Although it was not able to distinguish among the species of the B. cereus and B. subtilis groups, it should be considered a rapid and easy approach for the reliable identification of unknown Bacillus isolates or at least for the primary differentiation of Bacillus groups.

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Acknowledgements

We gratefully acknowledge Professor Alejandro Rooney from the Agricultural Research Service Culture Collection of USDA, for the species of B. subtilis group kindly provided. The authors thank Genome Sequencing Core-PDTIS/FIOCRUZ and INCQS Culture Collection for providing other the reference strains. We would also like to thank Dr. R. P. Vieira from the Molecular Biology Laboratory for the marine isolates and Adalberto Lamim da Silva from Informatics Center of INCQS for his contribution with the pictures assembly. This research was supported by INCQS/FIOCRUZ AND FAPERJ.

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Authors and Affiliations

  1. Department of Microbiology, National Institute of Quality Control in Health, FIOCRUZ, Avenida Brasil, 4365 - Manguinhos, Rio de Janeiro, RJ, CEP-21045-900, Brazil

    Catia A. C. Miranda & Maysa Mandetta Clementino

  2. Institute of Medical Biochemistry, CCS, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil

    Orlando B. Martins

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  1. Catia A. C. Miranda
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  2. Orlando B. Martins
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  3. Maysa Mandetta Clementino
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Correspondence to Maysa Mandetta Clementino.

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Miranda, C.A.C., Martins, O.B. & Clementino, M.M. Species-level identification of Bacillus strains isolates from marine sediments by conventional biochemical, 16S rRNA gene sequencing and inter-tRNA gene sequence lengths analysis. Antonie van Leeuwenhoek 93, 297–304 (2008). https://doi.org/10.1007/s10482-007-9204-0

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  • DOI: https://doi.org/10.1007/s10482-007-9204-0

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