VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

MPV005Iron-limitation triggers the virulence of Pseudomonasaeruginosa in urinary tract infectionsN. Rosin 1 , L. Jänsch 2 , M. Schobert 1 , D. Jahn 1 , P. Tielen* 11 Institute for Microbiology, University of Technology, Braunschweig,Germany2 Cellular Proteom Research, Helmholtz Center for Infection Research,Braunschweig, GermanyUrinary tract infections are one of the most common bacterial infections.Uncomplicated infections are mainly caused by Enterobacteriaceae.However, in case of complicated urinary tract infections Pseudomonasaeruginosa was identified as one of the most frequent pathogens. Theprogressive course of these infections is due to the remarkable ability of P.aeruginosa to adapt to hostile environments, its multifactorial virulence andits high intrinsic antibiotic resistance.An in vitro growth system mimicking the conditions in the urinary tract wasestablished to investigate the physiology of P. aeruginosa during urinarytract infections. Comprehensive transcriptome, proteome and metabolomeanalyses showed a general change in metabolic processes indicating that P.aeruginosa suffers from nutrient starvation and energy limitation. Moreover,in response to iron-limitation and osmotic stress a fine-tuned regulationcontrols the expression of several important virulence factors.In summary, the results indicate that the adaptative response of P.aeruginosa to the specific conditions in the urinary tract activates aregulatory network inducing the production of virulence factors.MPV006Metabolomic priming by a secreted fungal effectorA. Djamei*, K. Schipper, R. KahmannDepartment for Organismic Interactions, Max Planck Institute forTerrestrial Microbiology Marburg, GermanyA successful colonization of plants by pathogens requires active effectormediatedsuppression of defense responses. Here we show that thebiotrophic fungus Ustilago maydis secretes an enzymatically activechorismate mutase Cmu1. This enzyme is taken up locally by infected plantcells and then spreads to neighboring cells. Nonregulated enzymatic activityof the fungal chorismate mutase and interactions with cytoplasmic plantchorismate mutases are likely to be responsible for a re-channeling of theshikimate pathway. The comparison of the metabolomes of maize plantsinfected either with cmu1- deletion mutant or its progenitor strain showedsignificant changes in phenylpropanoid pathway derivates andphytohormone levels. Based on these findings, we propose a model in whichthe virulence factor Cmu1 actively reduces salicylic acid levels, therebyallowing the suppression of PAMP-triggered defense responses. Throughthis „metabolic priming”, the maize plant is prepared for a successfulinfection by Ustilago maydis. Our study describes a novel strategy for hostmodulation that might be used by a wide range of biotrophic plantpathogens.MPV007SACOL0731, a new regulatory link between centralcarbon metabolism and virulence determinantproduction in Staphylococcus aureusT. Hartmann 1 , R. Bertram 2 , W. Eisenreich 3 , B. Schulthess 4 , C. Wolz 5 ,M. Herrmann 1 , M. Bischoff* 11 Institute of Medical Microbiology and Hygine, Saarland UniversityHospital, Homburg/Saarbrücken, Germany2 Department of Microbial Genetics, Eberhard-Karls-University, Tübingen,Germany3 , Department of Biochemistry, Technical University Munich, München,Germany4 Institute of Medical Microbiology, University of Zurich, Zurich,Switzerland5 Institute for Medical Microbiology and Hygiene, University HospitalTübingen, Tübingen, Germanymember of the GalR-LacI repressor family. In Staphylococcus aureus, CcpAhas been shown to modulate the expression of metabolic genes and virulencedeterminants in response to glucose. A second regulator that links carbonmetabolism and virulence factor production in this organism is CodY, asensor of carbon and nitrogen availability that responds to intracellularconcentrations of branched-chain amino acids (BCAA) and GTP.Here we show that S. aureus produces a third regulatory molecule,SACOL0731 (a member of the LysR family of transcriptional regulatorswith homology to CitR of B. subtilis) that links central carbon metabolismwith virulence determinant production. By deleting this putative citRhomolog in S. aureus, we could show that the inactivation of this generesulted in a decreased citB (encoding the tricarboxylic acid [TCA] cycleenzyme aconitase) transcription, which was also illustrated by a stronglyreduced aconitase activity of the mutant under growth conditions that lackglucose. This regulatory effect was also confirmed by NMR-spectroscopystudies, which revealed an elevated citrate content in SACOL-0731 mutantcells. In line with previous findings showing that inactivation of the TCAcycle influences virulence determinant production of S. aureus, we foundthat the transcription of virulence factors such as capA (encoding capsularpolysaccharide synthesis enzyme A), hla (encoding α-hemolysin), and ofRNAIII, the effector molecule of the agr locus, were significantly affectedby the SACOL0731 mutation.MPV008Characterization of methionine auxotrophic clinicalPseudomonas aeruginosa isolatesA. Wesche* 1 , S. Thoma 1 , M. Hogardt 2 , E. Jordan 3 , D. Schomburg 3 ,M. Schobert 11 Institute for Microbiology, University of Technology, Braunschweig,Germany2 Max von Pettenkofer Institute, München, Germany3 Department of Bioinformatics and Biochemistry, University of Technology,Braunschweig, GermanyPatients with the genetic disorder cystic fibrosis (CF) suffer from increasedmucus production in the upper airways. This mucus is rich in nutrients ase.g. amino acids and is colonized by a heterogeneous microflora, whichcauses persistent infection. Infections with the opportunistic pathogen P.aeruginosa are associated with a poor prognosis due to the failure ofantibiotic treatment. P. aeruginosa colonizes CF mucus and adapts to the CFlung environment by mutation. Auxotrophic P. aeruginosa strains arefrequently isolated but their contribution to persistent infection is poorlyunderstood.Most auxotrophic strains require the amino acid methionine for growth.Interestingly, the methionine metabolism of P. aeruginosa is closelyconnected to the formation of the N-acyl-homoserine lactones (AHLs) thequorum sensing molecules.Here, we investigated and characterized 28 methionine auxotrophic P.aeruginosa isolates to elucidate the underlying adaptation strategies. Weidentified that methionine auxotrophy was caused by a mutation in the metFgene in 12 out of 28 clinical P. aeruginosa isolates. To elucidate thephenotype of a metF mutant, we constructed and characterized a knockoutmutant in P. aeruginosa PAO1. Growth experiments in M9 caseinate wereperformed and oxygen consumption during growth was determined for P.aeruginosa PAO1 wild type and the metF mutant. While we did not observeany growth differences between both strains, we noticed strongly reducedproduction of the virulence factors pyocyanin and the siderophore pyoverdinin the metF mutant. Since pyocyanin production is dependent on quorumsensing, we checked AHL production in the metF mutant strain.Unexpectedly, no difference to the PAO1 wild type strain was observed.This indicates that pyocyanin production is reduced in the metF mutantstrain by a quorum sensing independent pathway. Microarray andmetabolome analysis experiments are currently applied to elucidate therespective phenotype of the metF mutation.Carbon catabolite repression (CCR) in bacteria is a widespread, globalregulatory phenomenon that allows modulation of the expression of genesand operons involved in carbon utilization and metabolization in thepresence of preferred carbon source(s). In low-GC gram-positive bacteria,CCR is mediated mainly by the catabolite control protein A (CcpA), aspektrum | Tagungsband 2011