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The saltern-derived Paludifilum halophilum DSM 102817T is a new high-yield ectoines producer in minimal medium and under salt stress conditions

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Abstract

In the present study, the growth conditions and accumulation of ectoines (ectoine and hydroxyectoine) by Paludifilum halophilum DSM 102817T under salt stress conditions have been investigated. The productivity assay of this strain for ectoines revealed that the highest cellular content was reached in the minimal glucose sea water medium (SW-15) within 15% salinity. The addition of 0.1% (w/v) aspartic acid to the medium allowed an average of four times higher biomass production, and a dry mycelial biomass of 1.76 g L−1 was obtained after 6 days of growth in shake flasks at 40 °C and 200 rpm. Among the inorganic cations supplemented to the glucose SW-15 medium, the addition of 1 mM Fe2+ yielded the highest amount of mycelial biomass (3.45 g L−1) and total ectoines content (119 mg g−1), resulting in about 410 mg L−1 of products at the end of exponential growth phase. After 1 h of incubation in an osmotic downshock solution containing 2% NaCl, 70% of this content was released by the mycelium, and recovering cells maintained a high survival, with a maximal growth rate (µmax) of about 93% of the control population exposed to 15% NaCl. During growth at optimal salinity and temperature (15% NaCl and 40 °C), P. halophilum developed a compact and circular pellets that were easy to separate by simple decantation from both fermentation media and after hypoosmotic shock. Overall, the ectoines excreting P. halophilum could be a promising resource for ectoines production in a commercially valuable culture medium and at a large-scale fermentation process.

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References

  • Argandona M, Nieto JJ, Iglesias-Guerra F, Calderon MI, Garcia-Estepa R, Vargas C (2010) Interplay between iron homeostasis and the osmotic stress response in the halophilic bacterium Chromohalobacter salexigens. Appl Environ Microbiol 76:3575–3589

    CAS  PubMed  PubMed Central  Google Scholar 

  • Barth S, Huhn M, Matthey B, Klimka A, Galinski EA, Engert A (2000) Compatible-solute-supported periplasmic expression of functional recombinant proteins under stress conditions. Appl Environ Microbiol 66:1572–1579

    CAS  PubMed  PubMed Central  Google Scholar 

  • Ben Abdallah M, Karray F, Kallel N, Armougom F, Mhiri N, Quéméneur M, Cayol JL, Erauso G, Sayadi S (2018) Abundance and diversity of prokaryotes in ephemeral hypersaline lake Chott El Jerid using Illumina Miseq sequencing, DGGE and qPCR assays. Extremophiles 22:811–823

    CAS  PubMed  Google Scholar 

  • Borges N, Ramos A, Raven NDH, Sharp RJ (2002) Comparative study of the thermostabilizing properties of mannosylglycerate and other compatible solutes on model enzymes. Extremophiles 6:209–216

    CAS  PubMed  Google Scholar 

  • Boujelben I, Martínez-García M, van Pelt J, Maalej S (2015) Diversity of cultivable halophilic archaea and bacteria from superficial hypersaline sediments of Tunisian solar salterns. Antony van Leeuwenhoeck 106:675–692

    Google Scholar 

  • Brands S, Schein P, Castro-Ochoa KF, Galinski EA (2019) Hydroxyl radical scavenging of the compatible solute ectoine generates two N-acetimides. Arch Biochem Biophys 674:108097. https://doi.org/10.1016/j.abb.2019.108097

    Article  CAS  PubMed  Google Scholar 

  • Bünger J, Driller H (2004) Ectoine: an effective natural substance to prevent UVA-induced premature photoaging. Skin Pharmacol Physiol 17:232–237

    Google Scholar 

  • Bursy J, Pierik AJ, Pica N, Bremer E (2007) Osmotically induced synthesis of the compatible solute hydroxyectoine is mediated by an evolutionarily conserved ectoine hydroxylase. J Biol Chem 282:31147–31155

    CAS  PubMed  Google Scholar 

  • Bursy J, Kuhlmann AU, Pittelkow M, Hartmann H, Jebbar M, Pierik AJ, Bremer E (2008) Synthesis and uptake of the compatible solutes ectoine and 5-hydroxyectoine by Streptomyces coelicolor A3(2) in response to salt and heat stresses. Appl Environ Bacteriol 74:7286–7296

    CAS  Google Scholar 

  • Da Costa MS, Santos H, Galinski EA (1998) An overview of the role and diversity of compatible solutes in Bacteria and Archaea. Adv Biochem Eng Biotechnol 61:117–153

    PubMed  Google Scholar 

  • Deng Y, Huang L, Zhang C, Xie P, Cheng J, Wang X (2019) Physicochemical and functional properties of water soluble gum from Chaenomeles speciosa seeds. J Bioresour Bioprod 4:222–230

    CAS  Google Scholar 

  • Detkova EN, Boltyanskaya YV (2007) Osmoadaptation of haloalkaliphilic bacteria: role of osmoregulators and their possible practical application. Microbiology 76:511–522

    CAS  Google Scholar 

  • Eiberweiser A, Nazet A, Kruchinin SE, Fedotova MV, Buchner R (2015) Hydration and ion binding of the osmolyte ectoine. J Phys Chem B 119:15203–15211

    CAS  PubMed  Google Scholar 

  • Frikha-Dammak D, Fardeau ML, Cayol JL, Ben Fguira-Fourati L, Najeh S, Ollivier B, Maalej S (2016) Paludifilum halophilum gen. nov. sp. nov. a thermoactinomycete isolated from superficial sediment of a solar saltern. Int J Syst Evol Microbiol 66:1–8

    Google Scholar 

  • Galinski EA, Pfeiffer HP, Trüper HG (1985) 1,4,5,6-Tetrahydro-2-methyl-4-pyrimidinecarboxylic acid. A novel cyclic amino acid from halophilic phototrophic bacteria of the genus Ectothiorhodospira. Eur J Biochem 149:135–139

    CAS  PubMed  Google Scholar 

  • Graf R, Anzali S, Buenger J, Pfluecker F, Driller H (2008) The multifunctional role of ectoine as a natural cell protectant. Clin Dermatol 26:326–333

    PubMed  Google Scholar 

  • Guzmán H, Van-Thuoc D, Martín J, Hatti-Kaul R, Quillaguamán J (2009) A process for the production of ectoine and poly(3-hydroxybutyrate) by Halomonas boliviensis. Appl Microbiol Biotechnol 84:1069–1077

    PubMed  Google Scholar 

  • Han J, Gao Q-X, Zhang Y-G, Li L, Mohamad OAA, Narsing Rao MP, Xiao M, Hozzein WN, Alkhalifah DHM, Tao Y, Li W-J (2018) Transcriptomic and ectoine analysis of halotolerant Nocardiopsis gilva YIM 90087T under salt stress. Front Microbiol 9:618

    PubMed  PubMed Central  Google Scholar 

  • Harishchandra RK, Wulff S, Lentzen G, Neuhaus T, Galla HJ (2010) The effect of compatible solute ectoines on the structural organization of lipid monolayer and bilayer membranes. Biophys Chem 150:37–46. https://doi.org/10.1016/j.bpc.2010.02.007

    Article  CAS  PubMed  Google Scholar 

  • He YZ, Gong J, Yu HY, Tao Y, Zhang S, Dong ZY (2015) High production of ectoine from aspartate and glycerol by use of whole-cell biocatalysis in recombinant Escherichia coli. Microb Cell Fact 14:55

    PubMed  PubMed Central  Google Scholar 

  • Hille A, Neu TR, Hempel DC, Horn H (2005) Oxygen profiles and biomass distribution in biopellets of Aspergillus niger. Biotechnol Bioeng 92:614–623

    CAS  PubMed  Google Scholar 

  • Ignatova Z, Gierasch LM (2006) Inhibition of protein aggregation in vitro and in vivo by a natural osmoprotectant. Proc Natl Acad Sci USA 103:13357–13361

    CAS  PubMed  Google Scholar 

  • Ikeda H, Ishikawa J, Hanamoto A, Shinose M, Kikuchi H, Shiba T, Sakaki Y, Hattori M, Omura S (2003) Complete genome sequence and comparative analysis of the industrial microorganism Streptomyces avermitilis. Nat Biotechnol 21:526–531

    PubMed  Google Scholar 

  • Inbar L, Lapidot A (1988) The structure and biosynthesis of new tetrahydropyrimidine derivatives in actinomycin D producer Streptomyces parvulus. Use of 13C- and 15N-labeled L-glutamate and 13C and 15N NMR spectroscopy. J Biol Chem 263:16014–16022

    CAS  PubMed  Google Scholar 

  • Jorge CD, Borges N, Bagyan I, Bilstein A, Santos H (2016) Potential applications of stress solutes from extremophiles in protein folding diseases and healthcare. Extremophiles 20:251–259

    CAS  PubMed  Google Scholar 

  • Kempf B, Bremer E (1998) Uptake and synthesis of compatible solutes as microbial stress responses to high osmolality environments. Arch Microbiol 170:319–330

    CAS  PubMed  Google Scholar 

  • Klauck E, Typas A, Hengge R (2007) The sigmaS subunit of RNA polymerase as a signal integrator and network master regulator in the general stress response in Escherichia coli. Sci Prog 90:103–127

    CAS  PubMed  Google Scholar 

  • Kolp S, Pietsch M, Galinski EA, Gutschow M (2006) Compatible solutes as protectants for zymogens against proteolysis. Biochim Biophys Acta 1764:1234–1242. https://doi.org/10.1016/j.bbapap.2006.04.015

    Article  CAS  PubMed  Google Scholar 

  • Kunte HJ, Galinski EA, Trüper HG (1993) A modified FMOC-method for the detection of aminoacid type osmolytes and tetrahydropyrimidines (ectoines). J Microbiol Methods 17:129–136

    CAS  Google Scholar 

  • Kunte HJ, Lentzen G, Galinski E (2014) Industrial production of the cell protectant ectoine: protection, mechanisms, processes, and products. Curr Biotechnol 3:10–25

    CAS  Google Scholar 

  • Lentzen G, Schwarz T (2006) Extremolytes: natural compounds from extremophiles for versatile applications. Appl Microbiol Biotechnol 72:623–634

    CAS  PubMed  Google Scholar 

  • León MJ, Hoffmann T, Sánchez-Porro C, Heider J, Ventosa A, Bremer E (2018) Compatible solute synthesis and import by the moderate halophile Spiribacter salinus: physiology and genomics. Front Microbiol 9:108

    PubMed  PubMed Central  Google Scholar 

  • Liu KH, Ding XW, Narsing Rao MP, Zhang B, Zhang YG, Liu FH (2017) Morphological and transcriptomic analysis reveals the osmoadaptive response of endophytic fungus Aspergillus montevidensis ZYD4 to high salt stress. Front Microbiol 8:1789

    PubMed  PubMed Central  Google Scholar 

  • Malin G, Lapidot A (1996) Induction of synthesis of tetrahydropyrimidine derivatives in Streptomyces strains and their effect on Escherichia coli in response to osmotic and heat stress. J Bacteriol 178:385–395

    CAS  PubMed  PubMed Central  Google Scholar 

  • Motta A, Romano I, Gambacorta A (2004) Rapid and sensitive NMR method for osmolyte determination. J Microbiol Methods 58:289–294

    CAS  PubMed  Google Scholar 

  • O’Cleirigh C, Casey JT, Walsh PK, O’Shea DG (2005) Morphological engineering of Streptomyces hygroscopicus var.geldanus: regulation of pellet morphology through manipulation of broth viscosity. Appl Environ Microbiol 68:305–310

    Google Scholar 

  • Ono H, Sawada K, Khunajakr N, Tao T, Yamamoto M, Hiramoto M, Shinmyo A, Takano M, Murooka Y (1999) Characterization of biosynthetic enzymes for ectoine as a compatible solute in a moderately halophilic eubacterium, Halomonas elongata. J Bacteriol 181:91–99

    CAS  PubMed  PubMed Central  Google Scholar 

  • Onraedt AE, Walcarius BA, Soetaert WK, Vandamme EJ (2005) Optimization of ectoine synthesis through fed-batch fermentation of Brevibacterium epidermis. Biotechnol Prog 21:1206–1212

    CAS  PubMed  Google Scholar 

  • Oren A (2002) Intracellular salt concentration and ion metabolism in halophilic microorganisms. In: Seckbach J (ed) Halophilic microorganisms and their environments, 5th edn. Kluwer Academic Publishers, Dordrecht, pp 207–231

    Google Scholar 

  • Oren A (2013) Life at high salt concentrations, intracellular KCl concentrations, and acidic proteomes. Front Microbiol 4:315

    PubMed  PubMed Central  Google Scholar 

  • Öztürk HU, Sariyar Akbulut B, Ayan B, Poli A (2015) Moderately halophilic bacterium Halomonas sp. AAD12: a promising candidate as a hydroxyectoine producer. J Microb Biochem Technol 7:262–268

    Google Scholar 

  • Pastor JM, Salvador M, Argandona M, Bernal V, Reina-Bueno M, Csonka LN, Iborra JL, Vargas C, Nieto JJ, Canovas M (2010) Ectoines in cell stress protection: uses and biotechnological production. Biotechnol Adv 28:782–801

    CAS  PubMed  Google Scholar 

  • Prabhu J, Schauwecker F, Grammel N, Keller U, Bernhard M (2004) Functional expression of the ectoine hydroxylase gene (thpD) from Streptomyces chrysomallus in Halomonas elongata. Appl Environ Microbiol 70:3130–3132

    CAS  PubMed  PubMed Central  Google Scholar 

  • Rani A, Venkatesu P (2018) Changing relations between proteins and osmolytes: a choice of nature. Phys Chem Chem Phys 20:20315–20333

    CAS  PubMed  Google Scholar 

  • Reshetnikov AS, Khmelenina VN, Mustakhimov II, Trotsenko YA (2011) Genes and enzymes of ectoine biosynthesis in halotolerant methanotrophs. Methods Enzymol 495:15–30

    CAS  PubMed  Google Scholar 

  • Reuter K, Pittelkow M, Bursy J, Heine A, Craan T, Bremer E (2010) Synthesis of 5-hydroxyectoine from ectoine: crystal structure of the non-heme iron(II) and 2-oxoglutarate-dependent dioxygenase EctD. PLoS ONE 5(5):e10647. https://doi.org/10.1371/journal.pone.0010647

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Richter AA, Mais C-N, Czech L, Geyer K, Hoeppner A, Smits SHJ, Erb TJ, Bange G, Bremer E (2019) Biosynthesis of the stress-protectant and chemical chaperon ectoine: biochemistry of the transaminase EctB. Front Microbiol 10:2811. https://doi.org/10.3389/fmicb.2019.02811

    Article  PubMed  PubMed Central  Google Scholar 

  • Roberts MF (2005) Organic compatible solutes of halotolerant and halophilic microorganism. Saline Syst 1:5

    PubMed  PubMed Central  Google Scholar 

  • Sadeghi A, Soltani BM, Nekouei MK, Jouzani GS, Mirzaei HH, Sadeghizadeh M (2014) Diversity of the ectoines biosynthesis genes in the salt tolerant Streptomyces and evidence for inductive effect of ectoines on their accumulation. Microbiol Res 169:699–708

    CAS  PubMed  Google Scholar 

  • Sauer T, Galinski EA (1998) Bacterial milking: a novel bioprocess for production of compatible solutes. Biotechnol Bioeng 57:306–313

    CAS  PubMed  Google Scholar 

  • Schröter MA, Meyer S, Hahn MB, Solomun T, Sturm H, Kunte HJ (2017) Ectoine protects DNA from damage by ionizing radiation. Sci Rep 7:15272. https://doi.org/10.1038/s41598-017-15512-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sinha J, Bae JT, Park JP, Kim KH, Song CH, Yun JW (2001) Changes in morphology of Paecilomyces japonika and their effect on broth reology during production of exo-biopolymers. Appl Microbiol Biotechnol 56:88–92

    CAS  PubMed  Google Scholar 

  • Tao P, Li H, Yu Y, Gu J, Liu Y (2016) Ectoine and 5-hydroxyectoine accumulation in the halophile Virgibacillus halodenitrificans PDB-F2 in response to salt stress. Appl Microbiol Biotechnol 100:6779–6789

    CAS  PubMed  Google Scholar 

  • Tesche S, Rösemeier-Scheumann R, Lohr J, Hanke R, Büchs J, Krull R (2019) Salt-enhanced cultivation as a morphology engineering tool for filamentous actinomycetes: increased production of labyrinthopeptin A1 in Actinomadura namibiensis. Eng Life Sci 19:781–794

    CAS  PubMed  PubMed Central  Google Scholar 

  • Van Dissel D, Claessen D, van Wezel GP (2014) Morphogenesis of Streptomyces in submerged cultures. Adv Appl Microbiol 89:1–45

    PubMed  Google Scholar 

  • Van-Thuoc D, Guzmán H, Quillaguamán J, Hatti-Kaul R (2010) High productivity of ectoines by Halomonas boliviensis using a combined twostep fed-batch culture and milking process. J Biotechnol 147:46–51

    CAS  PubMed  Google Scholar 

  • Van-Thuoc D, Hashim SO, Hatti-Kaul R, Mamo G (2013) Ectoine mediated protection of enzyme from the effect of pH and temperature stress: a study using Bacillus halodurans xylanase as a model. Appl Microbiol Biotechnol 97:6271–6278

    CAS  PubMed  Google Scholar 

  • Van-Thuoc D, Thi Hien T, Sudesh K (2019) Identification and characterization of ectoine-producing bacteria isolated from Can Gio mangrove soil in Vietnam. Ann Microbiol 69:819–828

    CAS  Google Scholar 

  • Vargas C, Jebbar M, Carrasco R, Blanco C, Calderon M, Iglesias-guerra F, Nieto JJ (2005) Ectoines as compatible solutes and carbon and energy sources for the halophilic bacterium Chromohalobacter salexigens. J Appl Microbiol 100:98–107

    Google Scholar 

  • Vyrides I, Stuckey DC (2017) Compatible solute addition to biological systems treating waste/wastewater to counteract osmotic and other environmental stresses: a review. Crit Rev Biotechnol 37:865–879

    CAS  PubMed  Google Scholar 

  • Wei-Chuan C, Ching-Cha H, John Chi-Wei L, Yu-Kaung C, Li-Fen W, Yu-Hong W (2017) Production and characterization of ectoine using a moderately halophilic strain Halomonas salina BCRC17875. J Biosci Bioeng 125:1–7

    Google Scholar 

  • Werkhäuser N, Bilstein A, Sonnemann U (2014) Treatment of allergic rhinitis with ectoine containing nasal spray and eye drops in comparison with azelastine containing nasal spray and eye drops or with cromoglycic acid containing nasal spray. J Allergy. https://doi.org/10.1155/2014/176597

    Article  Google Scholar 

  • Widderich N, Kobus S, Höppner A, Riclea R, Seubert A, Dickschat JS, Heider J, Smits SHJ, Bremer E (2016) Biochemistry and crystal structure of ectoine synthase: a metal-containing member of the cupin superfamily. PLoS ONE. https://doi.org/10.1371/journal.pone.0151285

    Article  PubMed  PubMed Central  Google Scholar 

  • Wucherpfennig T, Kiep KA, Driouch H, Wittmann C, Krull R (2010) Morphology and rheology in filamentous cultivations. Adv Appl Micobiol 72:89–136

    CAS  Google Scholar 

  • Zhang Y, Li Y, Zhang Y, Wang Z, Zhao M, Su N et al (2016) Quantitative proteomics reveals membrane protein-mediated hypersaline sensitivity and adaptation in halophilic Nocardiopsis xinjiangensis. J Proteome Res 15:68–85

    CAS  PubMed  Google Scholar 

  • Zhao Q, Shannan L, Peiwen L, Simian S, Cuiqing M, Ping X, Haijun S, Chunyu Y (2019) High ectoine production by an engineered Halomonas hydrothermalis Y2 in a reduced salinity medium. Microb Cell Fact 18:184

    PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We are grateful to the General Direction of Scientific Research (DGRS) of Tunisia for financial support through the project PRF No PRF2019-D1P1.

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Authors and Affiliations

  1. Laboratoire de Biodiversité Marine et Environment (LR18ES/30), Université de Sfax, BP 1171, 3000, Sfax, Tunisia

    Houda Ayadi, Donyez Frikha-Dammak & Sami Maalej

  2. Laboratore de Chimie Organique (LR17ES/08), Unité des Substances Naturelles, Université de Sfax, BP 1171, 3000, Sfax, Tunisia

    Jawhar Fakhfakh & Noureddine Allouche

  3. Laboratore des Bioprocédés Environnementaux, Centre de Biotechnologie de Sfax, BP 1177, 3018, Sfax, Tunisia

    Mohamed Chamkha

  4. Unité de Valorisation des résultats de la Recherche, Centre de Biotechnologie de Sfax, BP 1177, 3018, Sfax, Tunisia

    Ilem Hassairi

  5. Center for Sustainable Development, College of Arts and Sciences, Qatar University, 2713, Doha, Qatar

    Sami Sayadi

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  1. Houda Ayadi
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Ayadi, H., Frikha-Dammak, D., Fakhfakh, J. et al. The saltern-derived Paludifilum halophilum DSM 102817T is a new high-yield ectoines producer in minimal medium and under salt stress conditions. 3 Biotech 10, 533 (2020). https://doi.org/10.1007/s13205-020-02512-x

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