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Applied and Environmental Microbiology, March 2007, p. 1992-2000, Vol. 73, No. 6
0099-2240/07/$08.00+0     doi:10.1128/AEM.02402-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Use of High-Affinity Cell Wall-Binding Domains of Bacteriophage Endolysins for Immobilization and Separation of Bacterial Cells{triangledown}

Jan W. Kretzer, Rainer Lehmann, Mathias Schmelcher, Manuel Banz, Kwang-Pyo Kim, Corinna Korn, and Martin J. Loessner*

Institute of Food Science and Nutrition, ETH Zurich, Schmelzbergstrasse 7, 8092 Zurich, Switzerland

Received 12 October 2006/ Accepted 21 January 2007

Immobilization and magnetic separation for specific enrichment of microbial cells, such as the pathogen Listeria monocytogenes, depends on the availability of suitable affinity molecules. We report here a novel concept for the immobilization and separation of bacterial cells by replacing antibodies with cell wall-binding domains (CBDs) of bacteriophage-encoded peptidoglycan hydrolases (endolysins). These polypeptide modules very specifically recognize and bind to ligands on the gram-positive cell wall with high affinity. With paramagnetic beads coated with recombinant Listeria phage endolysin-derived CBD molecules, more than 90% of the viable L. monocytogenes cells could be immobilized and recovered from diluted suspensions within 20 to 40 min. Recovery rates were similar for different species and serovars of Listeria and were not affected by the presence of other microorganisms. The CBD-based magnetic separation (CBD-MS) procedure was evaluated for capture and detection of L. monocytogenes from artificially and naturally contaminated food samples. The CBD separation method was shown to be superior to the established standard procedures; it required less time (48 h versus 96 h) and was the more sensitive method. Furthermore, the generalizability of the CBD-MS approach was demonstrated by using specific phage-encoded CBDs specifically recognizing Bacillus cereus and Clostridium perfringens cells, respectively. Altogether, CBD polypeptides represent novel and innovative tools for the binding and capture of bacterial cells, with many possible applications in microbiology and diagnostics.

* Corresponding author. Mailing address: Institute of Food Science and Nutrition, ETH Zurich, Schmelzbergstrasse 7, CH-8092 Zurich, Switzerland. Phone: 41 44 632 3335. Fax: 41 44 632 1266. E-mail: martin.loessner{at}ethz.ch.

{triangledown} Published ahead of print on 2 February 2007.

Applied and Environmental Microbiology, March 2007, p. 1992-2000, Vol. 73, No. 6
0099-2240/07/$08.00+0     doi:10.1128/AEM.02402-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Mayer, M. J., Narbad, A., Gasson, M. J. (2008). Molecular Characterization of a Clostridium difficile Bacteriophage and Its Cloned Biologically Active Endolysin. J. Bacteriol. 190: 6734-6740 [Abstract] [Full Text]  


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