RESUMO
The interest of the pharmaceutical industry in developing new antibiotics is decreasing, as established screening systems which identify compounds that kill or inhibit the growth of bacteria can no longer be used. Consequently, antimicrobial screening using classical minimum inhibitory concentration (MIC) measurements is becoming obsolete. The discovery of antimicrobial agents that specifically target a bacterial pathogen without affecting the host and its beneficial bacteria is a promising strategy. However, few host-microbe models are available for in vivo screening of novel antivirulence molecules. Here we designed high-throughput developmental assays in the social amoeba Dictyostelium discoideum to measure Pseudomonas aeruginosa virulence and to screen for novel antivirulence molecules without side effects to the host and its beneficial bacteria Klebsiella aerogenes. Thirty compounds were evaluated that had been previously selected by virtual screening for inhibitors of P. aeruginosa PAO1 polyphosphate kinase 1 (PaPPK1) and diverse compounds with combined PPK1 inhibitory and antivirulence activities were identified. This approach demonstrates that D. discoideum is a suitable surrogate host for preliminary high-throughput screening of antivirulence agents and that PPK1 is a suitable target for developing novel antivirulence compounds that can be further validated in mammalian models.
Assuntos
Anti-Infecciosos/isolamento & purificação , Dictyostelium/microbiologia , Avaliação Pré-Clínica de Medicamentos/métodos , Pseudomonas aeruginosa/efeitos dos fármacos , Anti-Infecciosos/farmacologia , Ensaios de Triagem em Larga Escala , Modelos Teóricos , Pseudomonas aeruginosa/patogenicidade , Pseudomonas aeruginosa/fisiologia , Virulência/efeitos dos fármacosRESUMO
BACKGROUND: Pseudomonas aeruginosa is known to be a multidrug resistant opportunistic pathogen. Particularly, P. aeruginosa PAO1 polyphosphate kinase mutant (ppk1) is deficient in motility, quorum sensing, biofilm formation and virulence. FINDINGS: By using Phenotypic Microarrays (PM) we analyzed near 2000 phenotypes of P. aeruginosa PAO1 polyP kinase mutants (ppk1 and ppk2). We found that both ppk mutants shared most of the phenotypic changes and interestingly many of them related to susceptibility toward numerous and different type of antibiotics such as Ciprofloxacin, Chloramphenicol and Rifampicin. CONCLUSIONS: Combining the fact that ppk1 mutants have reduced virulence and are more susceptible to antibiotics, polyP synthesis and particularly PPK1, is a good target for the design of molecules with anti-virulence and anti-persistence properties.
Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Análise em Microsséries/métodos , Mutação , Fosfotransferases (Aceptor do Grupo Fosfato)/genética , Pseudomonas aeruginosa/enzimologia , Antibacterianos/farmacologia , Cloranfenicol/farmacologia , Ciprofloxacina/farmacologia , Fenótipo , Polifosfatos/metabolismo , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/patogenicidade , Rifampina/farmacologia , Virulência/genéticaRESUMO
BACKGROUND: Pseudomonas aeruginosa is known to be a multidrug resistant opportunistic pathogen. Particularly, P. aeruginosa PAO1 polyphosphate kinase mutant (ppk1) is deficient in motility, quorum sensing, biofilm formation and virulence. FINDINGS: By using Phenotypic Microarrays (PM) we analyzed near 2000 phenotypes of P. aeruginosa PAO1 polyP kinase mutants (ppk1 and ppk2). We found that both ppk mutants shared most of the phenotypic changes and interestingly many of them related to susceptibility toward numerous and different type of antibiotics such as Ciprofloxacin, Chloramphenicol and Rifampicin. CONCLUSIONS: Combining the fact that ppk1 mutants have reduced virulence and are more susceptible to antibiotics, polyP synthesis and particularly PPK1, is a good target for the design of molecules with anti-virulence and anti-persistence properties.