Multidrug efflux pumps mediate resistance to antibiotics and other toxic compounds. We studied the role of AcrAB-TolC, the main efflux pump in Escherichia coli, in regulating gene expression. Deletion mutants, an acrABp-lacZ fusion and reverse transcription–real-time quantitative PCR experiments were used to study the role of AcrAB-TolC and metabolism in regulating gene expression of the acrAB operon and its transcriptional regulators. Deletion of the acrB gene increased the expression of the acrAB operon. A similar induction of acrAB was found when acrA or tolC was deleted, and when the pump function was inhibited using phenylalanine-arginine-β-naphthylamide. The induction of acrAB in the ΔacrB strain was totally (AcrR or SoxS) or partially (SoxR or MarA) prevented when the genes for these acrAB regulators were also deleted. The expression of soxS and marA, but not of acrR, was increased in the ΔacrB strain, which also showed altered expression of many other genes related to different cellular processes, including motility. Deletion of the metabolic genes entA and entE (enterobactin biosysnthesis), glpX (gluconeogenesis), cysH (cysteine biosynthesis) and purA (purine biosynthesis) also prevented activation of the acrAB promoter in the ΔacrB strain. Addition of the enterobactin biosynthesis intermediate metabolite 2,3-dihydroxybenzoate induced the expression of acrAB. These results together suggest a model in which the AcrAB-TolC pump effluxes cellular metabolites that are toxic and/or have a signalling role. If the pump is inactivated or inhibited, these metabolites would accumulate, inactivating AcrR and/or up-regulating soxS and marA expression, ultimately triggering the up-regulation of acrAB expression to restore homeostasis.

Multidrug resistance (MDR) in clinical isolates of Escherichia coli can be associated with overexpression of marA , a transcription factor that upregulates multidrug efflux and downregulates membrane permeability. Using random transposome mutagenesis, we found that many chromosomal genes and environmental stimuli affected MarA-mediated antibiotic resistance. Seven genes affected resistance mediated by MarA in an antibiotic-specific way; these were mostly genes encoding unrelated enzymes, transporters, and unknown proteins. Other genes affected MarA-mediated resistance to all antibiotics tested. These genes were acrA , acrB , and tolC (which encode the major MarA-regulated multidrug efflux pump AcrAB-TolC), crp , cyaA , hns , and pcnB (four genes involved in global regulation of gene expression), and the unknown gene damX . The last five genes affected MarA-mediated MDR by altering marA expression or MarA function specifically on acrA . These findings demonstrate that MarA-mediated MDR is regulated at multiple levels by different genes and stimuli, which makes it both complex and fine-tuned and interconnects it with global cell regulation and metabolism. Such a regulation could contribute to the adaptation and spread of MDR strains and may be targeted to treat antibiotic-resistant E. coli and related pathogens.

Carbapenems are β‐lactam antibiotics used in healthcare settings as last resort drugs to treat infections caused by antibiotic‐resistant bacteria. Carbapenem‐resistant bacteria are increasingly being isolated from healthcare facilities; however, little is known about their distribution or prevalence in the environment, especially in the United States, where their distribution in water environments from the West Coast has not been studied before. The aim of this study was to determine the prevalence of carbapenem‐resistant bacteria and carbapenemase genes in water bodies from the Los Angeles area (California, USA ). All samples that were analyzed contained carbapenem‐resistant bacteria with a frequency of between 0.1 and 324 carbapenem‐resistant cfu per 100 mls of water. We identified 76 carbapenem‐resistant or ‐intermediate isolates, most of which were also resistant to noncarbapenem antibiotics, as different strains of Enterobacter asburiae , Aeromonas veronii , Cupriavidus gilardii , Pseudomonas, and Stenotrophomonas species. Of them, 52 isolates were carbapenemase‐producers. Furthermore, PCR and sequence analysis to identify the carbapenemase gene of these carbapenemase‐producing isolates revealed that all Enterobacter asburiae isolates had a bla _{IMI ‐2} gene 100% identical to the reference sequence, and all Stenotrophomonas maltophlia isolates had a bla _L1 gene 83%–99% identical to the reference bla _L1 . Our findings indicate that water environments in Southern California are an important reservoir of bacteria‐resistant to carbapenems and other antibiotics, including bacteria carrying intrinsic and acquired carbapenemase genes.

MarA, a transcriptional regulator in Escherichia coli , affects functions such as multiple-antibiotic resistance (Mar) and virulence. Usually an activator, MarA is a repressor of hdeAB and other acid resistance genes. We found that, in wild-type cells grown in LB medium at pH 7.0 or pH 5.5, repression of hdeAB by MarA occurred only in stationary phase and was reduced in the absence of H-NS and GadE, the main regulators of hdeAB . Moreover, repression of hdeAB by MarA was greater in the absence of GadX or Lrp in exponential phase at pH 7.0 and in the absence of GadW or RpoS in stationary phase at pH 5.5. In turn, MarA enhanced repression of hdeAB by H-NS and hindered activation by GadE in stationary phase and also reduced the activity of GadX, GadW, RpoS, and Lrp on hdeAB under some conditions. As a result of its direct and indirect effects, overexpression of MarA prevented most of the induction of hdeAB expression as cells entered stationary phase and made the cells sevenfold more sensitive to acid challenge at pH 2.5. These findings show that repression of hdeAB by MarA depends on pH, growth phase, and other regulators of hdeAB and is associated with reduced resistance to acid conditions.