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Applied and Environmental Microbiology, March 2008, p. 1902-1908, Vol. 74, No. 6
0099-2240/08/$08.00+0     doi:10.1128/AEM.02539-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Emergence of Secretion-Defective Sublines of Pseudomonas aeruginosa PAO1 Resulting from Spontaneous Mutations in the vfr Global Regulatory Gene

Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Biologie Structurale et Microbiologie, Centre National de la Recherche Scientifique, 31 Chemin Joseph Aigier, 13402 Marseille Cedex 20, France,¹ Département de Microbiologie Fondamentale, Université de Lausanne, Bâtiment Biophore, CH-1015 Lausanne, Switzerland,² Institute of Infection, Immunity and Inflammation, University of Nottingham, University Park, Nottingham NG7 2RD, United Kingdom³

Received 10 November 2007/ Accepted 8 January 2008

Pseudomonas aeruginosa undergoes spontaneous mutation that impairs secretion of several extracellular enzymes during extended cultivation in vitro in rich media, as well as during long-term colonization of the cystic fibrosis lung. A frequent type of strong secretion deficiency is caused by inactivation of the quorum-sensing regulatory gene lasR. Here we analyzed a spontaneously emerging subline of strain PAO1 that exhibited moderate secretion deficiency and partial loss of quorum-sensing control. Using generalized transduction, we mapped the secretion defect to the vfr gene, which is known to control positively the expression of the lasR gene and type II secretion of several proteases. We confirmed this secretion defect by sequencing and complementation of the vfr mutation. In a reconstruction experiment conducted with a 1:1 mixture of wild-type strain PAO1 and a vfr mutant of PAO1, we observed that the vfr mutant had a selective advantage over the wild type after growth in static culture for 4 days. Under these conditions, spontaneous vfr emerged in a strain PAO1 population after four growth cycles, and these mutants accounted for more than 40% of the population after seven cycles. These results suggest that partial or complete loss of quorum sensing and secretion can be beneficial to P. aeruginosa under certain environmental conditions.

Published ahead of print on 18 January 2008.