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Manipulating Filamentous Fungus Chemical Phenotypes by Growth on Nutritional Arrays

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Fungal Secondary Metabolism

Part of the book series: Methods in Molecular Biology ((MIMB,volume 944))

Abstract

Methods for manipulating and fermenting microorganisms in multi-well plates offer unlimited possibilities for high-throughput parallel experimentation. Furthermore, bar-coded data tracking and downstream processing with modern liquid handling equipment reduce handling errors and are able to format microbial products for autosampler-equipped analytical instruments, e.g., HPLCs, mass spectrometers, and plate readers. An integrated system for high-throughput culturing of filamentous fungi replicating strains across many fermentation parameters, called nutritional arrays, was developed. It takes advantage of this equipment while addressing the age-old dilemma of how to manipulate fungal phenotypes to express a more complete spectrum of their secondary metabolites. Growth of any given strain in a well-designed nutritional array increases the chances of detecting a biologically active metabolite while reducing the manpower and materials needed for preparing individual fermentations and extracts. Fungi fermented in nutritional arrays are directly processed in a semi-automated fashion and the extracts prepared for bioassays and analytical chemistry. The necessary equipment, custom tools, and protocols to grow fungi in nutritional arrays are described along with examples of bioactive secondary metabolites discovered using this system.

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References

  1. Khaldi N, Seifuddin FT, Turner G, Haft D, Nierman WC, Wolfe KH, Fedorova ND (2010) SMURF: genomic mapping of fungal secondary metabolite clusters. Fungal Genet Biol 47:736–741

    Article  PubMed  CAS  Google Scholar 

  2. Brakhage AA, Schroeckh V (2010) Fungal secondary metabolites - strategies to activate silent gene clusters. Fungal Genet Biol 48:15–22

    Article  PubMed  Google Scholar 

  3. Hoffmeister D, Keller NP (2007) Natural products of filamentous fungi: enzymes, genes, and their regulation. Nat Prod Rep 24:393–516

    Article  PubMed  CAS  Google Scholar 

  4. Gross H (2007) Strategies to unravel the function of orphan biosynthesis pathways: recent examples and future prospects. Appl Microbiol Biotechnol 75:267–277

    Article  PubMed  CAS  Google Scholar 

  5. Yarbrough GG, Taylor DP, Rowlands RT, Crawford MS, Lasure LL (1993) Screening microbial metabolites for new drugs - theoretical and practical issues. J Antibiot 46:535–544

    Article  PubMed  CAS  Google Scholar 

  6. Höfs R, Walker M, Zeeck A (2000) Hexacyclinic acid, a polyketide from Streptomyces with a novel carbon skeleton. Angew Chem 39:3258–3261

    Article  Google Scholar 

  7. Bode HB, Bethe B, Höfs R, Zeeck A (2002) Big effects from small changes: possible ways to explore nature’s chemical diversity. Chembiochem 3:619–627

    Article  PubMed  CAS  Google Scholar 

  8. Scherlach K, Hertweck C (2006) Discovery of aspoquinolones A-D, prenylated quinoline-2-one alkaloids from Aspergillus nidulans, motivated by genome mining. Org Biomol Chem 4:3517–3520

    Article  PubMed  CAS  Google Scholar 

  9. Bills G, Platas G, Fillola A, Jiménez MR, Collado J, Vicente F, Martín J, González A, Bur-Zimmermann J, Tormo JR, Peláez F (2008) Enhancement of antibiotic and secondary metabolite detection from filamentous fungi by growth on nutritional arrays. J Appl Microbiol 104:1644–1658

    Article  PubMed  CAS  Google Scholar 

  10. Duetz W, Chase M, Bills G (2010) Miniaturization of fermentations. In: Demain A, Davies J, Baltz R (eds) Manual of industrial microbiology and biotechnology, 3rd edn. ASM, Washington, DC, pp 99–116

    Google Scholar 

  11. Duetz WA (2007) Microtiter plates as mini-bioreactors: miniaturization of fermentation methods. Trends Microbiol 15:469–475

    Article  PubMed  CAS  Google Scholar 

  12. Bills GF, Martín J, Collado J, Platas G, Overy D, Tormo JR, Vicente F, Verkleij G, Crous P (2009) Measuring the distribution and diversity of antibiosis and secondary metabolites in the filamentous fungi. Soc Ind Microbiol News 59:133–147

    Google Scholar 

  13. Roemer T, Xu DM, Singh SB, Parish CA, Harris G, Wang H, Davies JE, Bills GF (2011) Confronting the challenges of natural product-based antifungal discovery. Chem Biol 18:148–164

    Article  PubMed  CAS  Google Scholar 

  14. Vicente F, Basilio A, Platas G, Collado J, Bills GF, Del Val AG, Martin J, Tormo JR, Harris GH, Zink DL, Justice M, Kahn JN, Pelaez F (2009) Distribution of the antifungal agents sordarins across filamentous fungi. Mycol Res 113:754–770

    Article  PubMed  CAS  Google Scholar 

  15. Bills GF, Platas G, Overy D, Collado J, Fillola A, Jiménez MR, Martín J, del Val AG, Vicente F, Tormo JR, Peláez F, Calati K, Harris G, Parish C, Xu D, Roemer T (2009) Discovery of the parnafungins, antifungal metabolites that inhibit mRNA poly-adenylation, from the Fusarium larvarum complex and other hypocrealean fungi. Mycologia 101:445–469

    Article  Google Scholar 

  16. Peláez F, Collado J, Platas G, Overy DP, Martín J, Vicente F, González del Val A, Basilio A, de la Cruz M, Tormo JR, Fillola A, Arenal F, Villareal M, Rubio V, Baral HO, Galán R, Bills GF (2010) The phylogeny and intercontinental distribution of the pneumocandin-producing anamorphic fungus Glarea lozoyensis. Mycology 2:1–17

    Google Scholar 

  17. Ondeyka J, Harris G, Zink D, Basilio A, Vicente F, Bills G, Platas G, Collado J, González A, de la Cruz M, Martin J, Kahn JN, Galuska S, Giacobbe R, Abruzzo G, Hickey E, Liberator P, Jiang B, Xu DM, Roemer T, Singh SB (2009) Isolation, structure elucidation, and biological activity of virgineone from Lachnum virgineum using the genome-wide Candida albicans fitness fest. J Nat Prod 72:136–141

    Article  PubMed  CAS  Google Scholar 

  18. Herath K, Harris G, Jayasuriya H, Zink D, Smith S, Vicente F, Bills G, Collado J, Gonzalez A, Jiang B, Kahn JN, Galuska S, Giacobbe R, Abruzzo G, Hickey E, Liberator P, Xu DM, Roemer T, Singh SB (2009) Isolation, structure and biological activity of phomafungin, a cyclic lipodepsipeptide from a widespread tropical Phoma sp. Bioorg Med Chem 17:1361–1369

    Article  PubMed  CAS  Google Scholar 

  19. Xu D, Ondeyka J, Harris GH, Zink D, Nielsen-Kahn J, Wang H, Bills G, Platas G, Wang W, Szewczak AA, Liberator P, Roemer T, Singh SB (2011) Isolation, structure and biological activities of fellutamides C and D from an undescribed Metulocladosporiella (Chaetothyriales) using the genome-wide Candida albicans fitness test. J Nat Prod 74(8):1721–1730. doi:10.1021/np2001573

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

We would like to express our gratitude to all our former coworkers and colleagues who participated in the development, validation, and improvement of the nutritional array method.

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Correspondence to José R. Tormo .

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Tormo, J.R., Asensio, F.J., Bills, G.F. (2012). Manipulating Filamentous Fungus Chemical Phenotypes by Growth on Nutritional Arrays. In: Keller, N., Turner, G. (eds) Fungal Secondary Metabolism. Methods in Molecular Biology, vol 944. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-122-6_4

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  • DOI: https://doi.org/10.1007/978-1-62703-122-6_4

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-121-9

  • Online ISBN: 978-1-62703-122-6

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