Evolution of Pseudomonas aeruginosa virulence in infected patients revealed in a Dictyostelium discoideum host model E. Lelong, A. Marchetti, M. Simon,

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Evolution of Pseudomonas aeruginosa virulence in infected patients revealed in a Dictyostelium discoideum host model E. Lelong, A. Marchetti, M. Simon, J.L. Burns, C. van Delden, T. Köhler, P. Cosson Clinical Microbiology and Infection Volume 17, Issue 9, Pages 1415-1420 (September 2011) DOI: 10.1111/j.1469-0691.2010.03431.x Copyright © 2011 European Society of Clinical Infectious Diseases Terms and Conditions

FIG. 1 Measuring the virulence of Pseudomonas aeruginosa. The virulence of bacteria was determined by their ability to prevent growth of Dictyostelium discoideum. D. discoideum cells (10 000, 1 000, 100 or 10) were deposited on a lawn of bacteria in a 24-well plate. D. discoideum cells grew on avirulent bacteria and created a phagocytic plaque (white). To score the virulence, we determined, at days 4 and 10, the number of wells in which growth of D. discoideum was inhibited. The final virulence score was obtained by adding up the scores for days 4 and 10, yielding a number ranging from 0 (avirulent) to 8 (fully virulent). (a) PT5. (b) isolate 4A from patient 15 101. (c) PT462. (d) PT531 [4]. Clinical Microbiology and Infection 2011 17, 1415-1420DOI: (10.1111/j.1469-0691.2010.03431.x) Copyright © 2011 European Society of Clinical Infectious Diseases Terms and Conditions

FIG. 2 Virulence of Pseudomonas aeruginosa isolates from cystic fibrosis (CF) patients. The virulence of P. aeruginosa isolates collected from 16 different CF patients at different ages was determined as described in Fig. 1. Several individual clones were tested for each isolate (each dot represents one clone). Stars indicate isolates with lasR-inactivating mutations [2]. Clinical Microbiology and Infection 2011 17, 1415-1420DOI: (10.1111/j.1469-0691.2010.03431.x) Copyright © 2011 European Society of Clinical Infectious Diseases Terms and Conditions

FIG. 3 Virulence of Pseudomonas aeruginosa isolates from intubated patients. One hundred and sixteen P. aeruginosa isolates collected at daily intervals from ten intubated patients [3] were analysed as described in Fig. 1. (a) All isolates from patient 19 105 were fully virulent. A representative sample of all the isolates tested is shown. (b) For all patients listed, all isolates analysed were fully virulent, even though many had lasR inactivating mutations (stars). (c) Many isolates from patient 27 101 exhibited clones with decreased virulence. Six individual clones were tested for each isolate (each dot represents one clone). (d) Patient 15 101 was infected initially with strain OC2E, which was replaced after a few days with strain 239A. Isolates of strain OC2E were all virulent, whereas isolates of strain 239A exhibited heterogeneous virulence levels. (e) To evaluate the effect of lasR mutations on the virulence of 239A clones, individual colonies were analysed for the presence of lasR mutations, and their virulence was tested. *lasR mutated clone. ND, not determined; WT, wild type. Clinical Microbiology and Infection 2011 17, 1415-1420DOI: (10.1111/j.1469-0691.2010.03431.x) Copyright © 2011 European Society of Clinical Infectious Diseases Terms and Conditions