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Applied and Environmental Microbiology, January 2003, p. 49-55, Vol. 69, No. 1
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.1.49-55.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Recombinant Environmental Libraries Provide Access to Microbial Diversity for Drug Discovery from Natural Products

Sophie Courtois,1,2,{dagger} Carmela M. Cappellano,2 Maria Ball,3 Francois-Xavier Francou,3 Philippe Normand,1 Gérard Helynck,2 Asuncion Martinez,4 Steven J. Kolvek,4 Joern Hopke,4 Marcia S. Osburne,4* Paul R. August,4 Renaud Nalin,1,{ddagger} Michel Guérineau,3 Pascale Jeannin,2 Pascal Simonet,1 and Jean-Luc Pernodet3

Laboratoire d'Ecologie Microbienne du Sol, UMR CNRS 5557, Université Claude Bernard Lyon 1, 69622 Villeurbanne Cedex,1 Aventis Pharma, Centre de Recherche de Vitry-Alfortville, 94403 Vitry sur Seine Cedex,2 Institut de Génétique et Microbiologie, UMR CNRS 8621, Université Paris Sud XI, 91405 Orsay Cedex, France,3 Aventis Pharmaceuticals Inc., Cambridge Genomics Center, Cambridge, Massachusetts 021394

Received 10 July 2002/ Accepted 1 October 2002

To further explore possible avenues for accessing microbial biodiversity for drug discovery from natural products, we constructed and screened a 5,000-clone "shotgun" environmental DNA library by using an Escherichia coli-Streptomyces lividans shuttle cosmid vector and DNA inserts from microbes derived directly (without cultivation) from soil. The library was analyzed by several means to assess diversity, genetic content, and expression of heterologous genes in both expression hosts. We found that the phylogenetic content of the DNA library was extremely diverse, representing mostly microorganisms that have not been described previously. The library was screened by PCR for sequences similar to parts of type I polyketide synthase genes and tested for the expression of new molecules by screening of live colonies and cell extracts. The results revealed new polyketide synthase genes in at least eight clones. In addition, at least five additional clones were confirmed by high-pressure liquid chromatography analysis and/or biological activity to produce heterologous molecules. These data reinforce the idea that exploiting previously unknown or uncultivated microorganisms for the discovery of novel natural products has potential value and, most importantly, suggest a strategy for developing this technology into a realistic and effective drug discovery tool.

* Corresponding author. Mailing address: Aventis Cambridge Genomics Center, 26 Landsdowne St., Cambridge, MA 02139. Phone: (617) 768-4101. Fax: (617) 374-8811. E-mail: drothstein{at}rcn.com.

{dagger} Present address: ONDEO Services, Centre International de Recherche sur l'Eau et l'Environnement, 78 230 Le Pecq, France.

{ddagger} Present address: LIBRAGEN, 69622 Villeurbanne Cedex 2, France.

Applied and Environmental Microbiology, January 2003, p. 49-55, Vol. 69, No. 1
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.1.49-55.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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