CzcR-CzcS, a Two-component System Involved in Heavy Metal and Carbapenem Resistance in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is an environmental bacterium involved in mineralization of organic matter. It is also an opportunistic pathogen able to cause serious infections in immunocompromised hosts. As such, it is exposed to xenobiotics including solvents, heavy metals, and antimicrobials. We studied the response of P. aeruginosa upon exposure to heavy metals or antibiotics to investigate whether common regulatory mechanisms govern resistance to both types of compounds. We showed that sublethal zinc concentrations induced resistance to zinc, cadmium, and cobalt, while lethal zinc concentrations selected mutants constitutively resistant to these heavy metals. Both zinc-induced and stable zinc-resistant strains were also resistant to the carbapenem antibiotic imipenem. On the other hand, only 20% of clones selected on imipenem were also resistant to zinc. Heavy metal resistance in the mutants could be correlated by quantitative real time PCR with increased expression of the heavy metal efflux pump CzcCBA and its cognate two-component regulator genes czcR-czcS. Western blot analysis revealed reduced expression of the basic amino acid and carbapenem-specific OprD porin in all imipenem-resistant mutants. Sequencing of the czcR-czcS DNA region in eight independent zincand imipenem-resistant mutants revealed the presence of the same V194L mutation in the CzcS sensor protein. Overexpression in a susceptible wild type strain of the mutated CzsS protein, but not of the wild type form, resulted in decreased oprD and increased czcC expression. We further show that zinc is released from latex urinary catheters into urine in amounts sufficient to induce carbapenem resistance in P. aeruginosa, possibly compromising treatment of urinary tract infections by this class of antibiotics.Pseudomonas aeruginosa is a Gram-negative bacterium thriving in environments polluted with organic matter. It is also an opportunistic pathogen frequently encountered in the hospital, causing morbidity and mortality in immunocompromised and cystic fibrosis patients (1). P. aeruginosa is characterized by an intrinsically high level of resistance to xenobiotics including antimicrobial agents, solvents, and heavy metals (2), which can be accounted for by a combination of its low outer membrane permeability and the presence of multiple efflux pumps (3). These pumps belong to the resistance, nodulation, cell division (RND) 1 transporter family, present in many Gramnegative bacteria (4). To extrude substrates from the cytoplasm across the two membranes, these systems are composed of a proton antiporter located in the cytoplasmic membrane, a membrane fusion protein spanning the periplasmic space, and an outer membrane protein (5). Members of the RND family, namely the Mex pumps, have recently gained increasing interest. In particular, the constitutively expressed MexAB-OprM (6, 7) and the inducible MexXY (8) efflux pumps endow the PAO1 reference strain and other clinical isolates (9) with a natural resistance to a wide range of antimicrobial agents. Proton-dr...

Multidrug resistant Acinetobacter baumannii--the role of AdeABC (RND family) efflux pump in resistance to antibiotics.

“…This system is associated with different pumps of RND family appearing in Gram-negative bacteria [8,25,60,81,86,94], among others AdeABC of Acinetobacter baumannii. Level of the expression of the pump, regulated by such a system, is dependent on the presence of the stimulus (substrate) in environment of the cell, which activating the pump for removing substrate outside the cell.…”

Pseudomonas aeruginosa PAO1 Resistance to Zinc Pyrithione: Phenotypic Changes Suggest the Involvement of Efflux Pumps

“…ions seems to have induced it. However, the downregulation of OprD protein that was mentioned by Perron et al [21] was not observed. Furthermore, the CzcCBA efflux system in P. aeruginosa pumps Zn 2?…”

“…Zinc resistance in P. aeruginosa was attributed to the heavy metal efflux pump CzcCBA, which is under the control of the two-component regulatory system CzcRS [4,21]. We do not rule out the involvement of the CzcCBA system because the presence of Zn 2?…”

“…Scientists believe that bacteria cannot develop resistance to such a powerful biocide with multiple antimicrobial mechanisms [10] Previously, it has been shown that resistance of Pseudomonas aeruginosa strain PAO1 to ZnPT could be induced, and that this resistance was accompanied by loss of an outer membrane protein and increased insusceptibility toward other biocides [1]. Other authors have discussed the involvement of the CzcCBA efflux system in resistance of P. aeruginosa to zinc and its relation to imipenem resistance [4,21].…”

“…Efflux systems use energy for the antiport of harmful substances, including antibiotics, numerous dyes, detergents, inhibitors, organic solvents, and biocides, out of the cell [2,21]. Attacking the energy source will inhibit the efflux pump and allow the accumulation of harmful substances inside the bacteria to a toxic level that will reduce its tolerance to such substances.…”