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Antagonistic interactions and production of halocin antimicrobial peptides among extremely halophilic prokaryotes isolated from the solar saltern of Sfax, Tunisia

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Abstract

Thirty-five extremely halophilic microbial strains isolated from crystallizer (TS18) and non-crystallizer (M1) ponds in the Sfax solar saltern in Tunisia were examined for their ability to exert antimicrobial activity. Antagonistic assays resulted in the selection of eleven strains that displayed such antimicrobial activity and they were further characterized. Three cases of cross-domain inhibition (archaea/bacteria or bacteria/archaea) were observed. Four archaeal strains exerted antimicrobial activity against several other strains. Three strains, for which several lines of evidence suggested the antimicrobial activity was, at least in part, due to peptide/protein agents (Halobacterium salinarum ETD5, Hbt. salinarum ETD8, and Haloterrigena thermotolerans SS1R12), were studied further. Optimal culture conditions for growth and antimicrobial production were determined. Using DNA amplification with specific primers, sequencing and RT-PCR analysis, Hbt. salinarum ETD5 and Hbt. salinarum ETD8 were shown to encode and express halocin S8, a hydrophobic antimicrobial peptide targeting halophilic archaea. Although the gene encoding halocin H4 was amplified from the genome of Htg. thermotolerans SS1R12, no transcript could be detected and the antimicrobial activity was most likely due to multiple antimicrobial compounds. This is also the first report that points to four different strains isolated from different geographical locations with the capacity to produce identical halocin S8 proteins.

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Abbreviations

OD:

Optical density

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Acknowledgments

This study was supported by the Tunisian Ministry of Scientific Research and the French Ministry of Foreign Affairs through a joint project PHC-Utique No 13G0833, the University of Sfax (Tunisia) and the National Museum of Natural History in Paris (France). We are grateful to the Cotusal salt company in Sfax for permission to access and sample the solar saltern.

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Authors and Affiliations

  1. Unité Biodiversité et Ecosystèmes Aquatiques Environnementaux (UR11ES/72), Faculté des Sciences de Sfax, Université de Sfax, BP 1171, 3000, Sfax, Tunisia

    Fadoua Ghanmi, Ines Boujelben, Doniez Frikha, Habib Ayadi & Sami Maalej

  2. Molécules de Communication et Adaptation des Microorganismes (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum national d’Histoire naturelle, Centre national de la Recherche scientifique (CNRS), CP 54, 57 rue Cuvier, 75005, Paris, France

    Fadoua Ghanmi, Alyssa Carré-Mlouka, Manon Vandervennet, Jean Peduzzi & Sylvie Rebuffat

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  1. Fadoua Ghanmi
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  2. Alyssa Carré-Mlouka
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Correspondence to Alyssa Carré-Mlouka.

Additional information

Communicated by A. Oren.

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792_2016_827_MOESM1_ESM.pptx

Supplementary Figure S1: Amplification of halocin genes with halS8-specific primers halS8-F1 and halS8-R1 in Hbt. salinarum ETD5 and Hbt. salinarum ETD8 (a) and with halH4-specific primers halH4-F1 and halH4-R1in Htg. thermotolerans SS1R12 (b). a M Marker ExactLadder® DNA PreMix 2 Log (Ozyme, France), 1 Hbt. salinarum ETD5, 2 Hbt. salinarum ETD8. b M Marker ExactLadder® DNA PreMix 2 Log (Ozyme, France), 1-2 Htg. thermotolerans SS1R12, 3 Hfx. mediterranei DSM1411. (PPTX 105 kb)

792_2016_827_MOESM2_ESM.docx

Supplementary Figure S2: Alignments of the predicted amino acid sequences deduced from the nucleotide sequences of the halS8 gene from the uncharacterized halophilic archaea S8a (Genbank accession no AF276080, Price and Shand 2000), Halobacterium sp. GN 101 (Genbank accession no EU080936, Besse et al 2015), Hbt. salinarum ETD5 (Genbank accession no KT783468, this study) and Hbt. salinarum ETD8 (Genbank accession no KR611166, this study). The amino acid substitutions are indicated in shaded light gray. The sequence of the mature HalS8 peptide is underlined (Price and Shand, 2000 (DOCX 14 kb)

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Supplementary material 4 (DOCX 15 kb)

Supplementary material 5 (DOCX 15 kb)

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Ghanmi, F., Carré-Mlouka, A., Vandervennet, M. et al. Antagonistic interactions and production of halocin antimicrobial peptides among extremely halophilic prokaryotes isolated from the solar saltern of Sfax, Tunisia. Extremophiles 20, 363–374 (2016). https://doi.org/10.1007/s00792-016-0827-9

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