Identification of a ferritin-like protein of Listeria monocytogenes as a mediator of β-lactam tolerance and innate resistance to cephalosporins

Abstract

BackgroundThe food-borne pathogen Listeria monocytogenes is the causative agent of listeriosis. The β-lactam antibiotics penicillin G and ampicillin are the current drugs of choice for the treatment of listerial infections. While isolates of L. monocytogenes are susceptible to these antibiotics, their action is only bacteriostatic and consequently, this bacterium is regarded as tolerant to β-lactams. In addition, L. monocytogenes has a high level of innate resistance to the cephalosporin family of β-lactams frequently used to treat sepsis of unknown etiology. Given the high mortality rate of listeriosis despite rational antibiotic therapy, it is important to identify genes that play a role in the susceptibility and tolerance of L. monocytogenes to β-lactams.ResultsThe hly-based promoter trap system was applied to identify penicillin G-inducible genes of L. monocytogenes. The results of reporter system studies, verified by transcriptional analysis, identified ten penicillin G-inducible genes. The contribution of three of these genes, encoding a ferritin-like protein (fri), a two-component phosphate-response regulator (phoP) and an AraC/XylS family transcriptional regulator (axyR), to the susceptibility and tolerance of L. monocytogenes to β-lactams was examined by analysis of nonpolar deletion mutants. The absence of PhoP or AxyR resulted in more rapid growth of the strains in the presence of sublethal concentration of β-lactams, but had no effect on the MIC values or the ability to survive a lethal dose of these antibiotics. However, the Δfri strain showed impaired growth in the presence of sublethal concentrations of penicillin G and ampicillin and a significantly reduced ability to survive lethal concentrations of these β-lactams. A lack of Fri also caused a 2-fold increase in the sensitivity of L. monocytogenes to cefalotin and cephradine.ConclusionsThe present study has identified Fri as an important mediator of β-lactam tolerance and innate resistance to cephalosporins in L. monocytogenes. PhoP and AxyR are probably involved in transmitting signals to adjust the rate of growth of L. monocytogenes under β-lactam pressure, but these regulators do not play a significant role in susceptibility and tolerance to this class of antibiotics.

Critical Role of a Ferritin-Like Protein in the Control of Listeria monocytogenes Cell Envelope Structure and Stability under β-lactam Pressure

“…It has recently been shown that the Fri protein of L. monocytogenes plays a crucial role in cell death caused by β-lactam antibiotics [20]. Detailed studies were undertaken to define the involvement of Fri in the maintenance of L. monocytogenes cell envelope structure and stability under β-lactam pressure.…”

“…In this study, we compared the properties of wild-type strain EGD and the Δ fri mutant grown in both the absence and presence of 0.09 µg/ml of penicillin G (corresponds to 0.75 of the MIC value). These β-lactam stress conditions were previously shown to permit efficient growth of both strains, but caused slight growth retardation of strain Δ fri compared to the wild type [20]. To examine the role of Fri in maintaining the integrity of the L. monocytogenes cell envelope under β-lactam pressure, the susceptibility of the studied strains to lysis induced by incubation with the cell wall hydrolase lysozyme was assessed (Figure 1).…”

“…The L. monocytogenes strains used in all experiments were cultured as described previously [20]. Briefly, the strains were grown overnight in BHI medium at 37°C with shaking.…”

“…It was recently shown that Fri, a ferritin-like protein, plays a crucial role in the tolerance of L. monocytogenes to β-lactam antibiotics [20]. Fri of L. monocytogenes belongs to the Dps ( D NA-binding p roteins from s tarved cells) family of proteins, which play an important role in counteracting the adverse effects of iron under aerobic conditions.…”

“…The ferritin-like protein of L. monocytogenes contributes to virulence and plays a role in protection against multiple stresses including oxidative, iron-starvation, cold- and heat-shock [32], [37], [38]. In addition, the deletion of L. monocytogenes fri was shown to abolish the innate tolerance of this bacterium to β-lactam antibiotics, which was accompanied by visible lysis of the cells [20], [39]. These observations led us to hypothesize that the role of Fri protein in the tolerance of L. monocytogenes to β-lactams might be associated with the regulation of bacterial cell envelope integrity.…”