RESUMEN
We designed and constructed an antibiotic screening system by using antibiotic responsive genes as reporters. Plasmid pCS26 carrying a promoterless luminescence reporter, luxCDABE, was used as the vector and the promoter regions of antibiotic responsive genes/operons from Escherichia coli were cloned upstream of the lux reporter to form the first part of the screening reporter array. Random promoter library of Salmonella enterica and Pseudomonas aeruginosa were screened for antibiotic responsive clones which consist of the second part of the screening array. The selected final reporter array responded to different antibiotics in distinct patterns and enabled in vivo high-throughput screening for antibiotics. Unknown antibiotics could, in general, be classified by analyzing the response patterns. This screening system is both sensitive and efficient and should prove to be a useful tool for screening new antibiotic compounds.
Asunto(s)
Antibacterianos , Farmacología , Evaluación Preclínica de Medicamentos , Métodos , Regulación Bacteriana de la Expresión Génica , Regiones Promotoras Genéticas , Genética , Pseudomonas aeruginosa , Genética , Salmonella enterica , GenéticaRESUMEN
Quorum sensing is an important gene regulatory mechanism in Pseudomonas aeruginosa and controls the expression of numerous virulence factors. We designed and constructed a screening system for quorum-sensing inhibitors. We developed the system by using the lasI and rhlA promoters fused with promoterless sacB as reporters. Using this system we screened a number of Chinese herb extracts, and identified three herb extracts containing inhibitors to the quorum-sensing system and to its regulated genes. The screening system developed was highly efficient and sensitive. It could serve as a useful tool to identify herb compounds that block infections but unlikely render antibiotic resistance in pathogens.