RESUMO
The quorum-sensing (QS) system of Pseudomonas aeruginosa dominates the pathogenicity of the acute or chronic infection process. Hence, curbing the pathogenicity of P. aeruginosa by targeting QS system is an ideal strategy. This study aims to identify potential anti-virulence compounds that can effectively disrupt the QS system of P. aeruginosa using a combination of virtual screening and experimental validation techniques. We explored inhibitory effect of isovanillin obtained by virtual screening on P. aeruginosa QS regulated virulence factors extracellular protease, biofilm, and pyocyanin. Results displayed that isovanillin could inhibit the virulence phenotypes regulated by the las- and pqs-QS systems of P. aeruginosa. The synthesis of extracellular proteases, pyocyanin, and biofilm formation by P. aeruginosa were dramatically inhibited by sub-MICs doses of isovanillin. The results of RNA sequencing and quantitative PCR revealed that the QS-activated genes down-regulated by subinhibitory isovanillin in the transcriptional evels. Furthermore, the presence of isovanillin increased the susceptibility of drug-resistant P. aeruginosa to kanamycin, meropenem, and polymyxin B. Treatment of isovanillin as a monotherapy significantly decreased the mortality of C. elegans in P. aeruginosa PAO1 or UCBPP-PA14 (PA14) infection. Our study reported the anti-virulence activity of isovanillin against P. aeruginosa, and provided a structural foundation for developing anti-virulence drugs targeting the QS system of P. aeruginosa.
Assuntos
Antibacterianos , Biofilmes , Caenorhabditis elegans , Testes de Sensibilidade Microbiana , Pseudomonas aeruginosa , Piocianina , Percepção de Quorum , Fatores de Virulência , Percepção de Quorum/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/patogenicidade , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Virulência/efeitos dos fármacos , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Animais , Antibacterianos/farmacologia , Caenorhabditis elegans/microbiologia , Caenorhabditis elegans/efeitos dos fármacos , Piocianina/metabolismo , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Infecções por Pseudomonas/microbiologia , Peptídeo Hidrolases/metabolismo , Peptídeo Hidrolases/genéticaRESUMO
Colonization of Pseudomonas aeruginosa in the lung environments frequently leads to the enrichment of strains displaying enhanced antibiotic resistance and reduced production of quorum-sensing (QS) controlled products. However, the relationship between the emergence of QS deficient variants and antibiotic resistance remains less understood. In this study, 67 P. aeruginosa strains were isolated from the lungs of 14 patients with chronic obstructive pulmonary disease, followed by determining their genetic relationship, QS-related phenotypes and resistance to commonly used antibiotics. The integrity of P. aeruginosa QS system was checked by DNA sequencing. The relationship between the QS system and antibiotic resistance was then assessed by correlation analyses. The function of the LasR protein and bacterial virulence were evaluated through homology modeling and nematode-infection assay. The influence of antibiotic on the development of extracellular protease production ability of P. aeruginosa was tested by an evolutionary experiment. The results showed that P. aeruginosa clinical strains displayed abundant diversity in phenotype and genotype. The production of extracellular proteases was significantly negatively correlated with antibiotic resistance. The strains with enhanced antibiotic resistance also showed a notable overlap with the mutation of lasR gene, which is the core regulatory gene of P. aeruginosa QS system. Molecular docking and Caenorhabditis elegans infection assays further suggested that P. aeruginosa with impaired LasR protein could also have varying pathogenicity. Moreover, in vitro evolution experiments demonstrated that antibiotic-mediated selective pressure, particularly from Levofloxacin contributed to the emergence of extracellular protease-negative strains. Therefore, this study provides evidence for the connection of P. aeruginosa QS system and antibiotic resistance, and holds significance for developing targeted strategies to address antibiotic resistance and improving the management of antibiotic-resistant infections in chronic respiratory diseases.