RamA confers multidrug resistance in Salmonella enterica via increased expression of acrB, which is inhibited by chlorpromazine.
Salmonella enterica serovar Typhimurium SL1344, in which efflux pump genes (acrB, acrD, acrF, tolC) or regulatory genes thereof (marA, soxS, ramA) were inactivated, was grown in the presence of 240 antimicrobial and nonantimicrobial agents in the Biolog Phenotype MicroArray. Mutants lacking tolC, acrB, and ramA grew significantly worse than other mutants in the presence of 48 agents (some of which have not previously been identified as substrates of AcrAB-TolC) and particularly poorly in the presence of phenothiazines, which are human antipsychotics. MIC testing revealed that the phenothiazine chlorpromazine had antimicrobial activity and synergized with common antibiotics against different Salmonella serovars and SL1344. Chlorpromazine increased the intracellular accumulation of ethidium bromide, which was ablated in mutants lacking acrB, suggesting an interaction with AcrB. High-level but not low-level overexpression of ramA increased the expression of acrB; conferred resistance to chloramphenicol, tetracycline, nalidixic acid, and triclosan and organic solvent tolerance; and increased the amount of ethidium bromide accumulated. Chlorpromazine induced the modest overproduction of ramA but repressed acrB. These data suggest that phenothiazines are not efflux pump inhibitors but influence gene expression, including that of acrB, which confers the synergy with antimicrobials observed.
Bailey
Andrew M
AM
Antimicrobial Agents Research Group, Division of Immunity and Infection, University of Birmingham, Birmingham B15 2TT, United Kingdom.
Paulsen
Ian T
IT
Piddock
Laura J V
LJ
D000900:Anti-Bacterial Agents / Q000008:administration & dosage; D014150:Antipsychotic Agents / Q000008:administration & dosage / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D002746:Chlorpromazine / Q000008:administration & dosage / Q000494:pharmacology*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004357:Drug Synergism; D004996:Ethidium / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D005798:Genes, Bacterial; D006801:Humans; D026901:Membrane Transport Proteins / Q000235:genetics*; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics*; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D010640:Phenothiazines / Q000008:administration & dosage / Q000494:pharmacology; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics*
D000900:Anti-Bacterial Agents; D014150:Antipsychotic Agents; D001426:Bacterial Proteins; D002746:Chlorpromazine; D017931:DNA Primers; D004269:DNA, Bacterial; D004996:Ethidium; D026901:Membrane Transport Proteins; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D010640:Phenothiazines; D015534:Trans-Activators; C474926:AcrB protein, Salmonella enterica; C531999:RamA protein, Salmonella typhimurium