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The Scientific World Journal
Volume 2012, Article ID 503269, 9 pages
http://dx.doi.org/10.1100/2012/503269
Research Article

Expanding the Use of a Fluorogenic Method to Determine Activity and Mode of Action of Bacillus thuringiensis Bacteriocins Against Gram-Positive and Gram-Negative Bacteria

1Escuela de Ciencias Biológicas, Universidad Autónoma de Coahuila, 27440 Torreón, COAH, Mexico
2División Ciencias de la Vida, Departamento de Alimentos, Universidad de Guanajuato Campus Irapuato-Salamanca, 36500 Irapuato, GTO, Mexico
3Department of Natural and Mathematical Sciences, California Baptist University, 8432 Magnolia Avenue, Riverside, CA 92504, USA
4Department of Entomology, University of California, Riverside, CA 92521, USA

Received 16 April 2012; Accepted 8 May 2012

Academic Editors: S. Fuchs, K. Hong, E. Parrilli, and L. Ramirez

Copyright © 2012 Norma M. de la Fuente-Salcido et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Previously we described a rapid fluorogenic method to measure the activity of five bacteriocins produced by Mexican strains of Bacillus thuringiensis against B. cereus 183. Here we standardize this method to efficiently determine the activity of bacteriocins against both Gram-positive and Gram-negative bacteria. It was determined that the crucial parameter required to obtain reproducible results was the number of cells used in the assay, that is, ~4 × 108 cell/mL and ~7 × 108 cell/mL, respectively, for target Gram-positive and Gram-negative bacteria. Comparative analyses of the fluorogenic and traditional well-diffusion assays showed correlation coefficients of 0.88 to 0.99 and 0.83 to 0.99, respectively, for Gram-positive and Gram-negative bacteria. The fluorogenic method demonstrated that the five bacteriocins of B. thuringiensis have bacteriolytic and bacteriostatic activities against all microorganisms tested, including clinically significant bacteria such as Listeria monocytogenes, Proteus vulgaris, and Shigella flexneri reported previously to be resistant to the antimicrobials as determined using the well-diffusion protocol. These results demonstrate that the fluorogenic assay is a more sensitive, reliable, and rapid method when compared with the well-diffusion method and can easily be adapted in screening protocols for bacteriocin production by other microorganisms.