Beta-lactamase Inhibitor Tazobactam Research Articles

LC-MS/MS-based multiplex antibacterial platform for therapeutic drug monitoring in intensive care unit patients.

Empirically prescribed standard dosing regimens of antibacterial agents may result in insufficient or excess plasma concentrations with persistently poor clinical outcomes, especially for patients in intensive care units (ICUs). Therapeutic drug monitoring (TDM) of antibacterial agents can guide dose adjustments to benefit patients. In this study, we developed a robust, sensitive, and simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) platform for the quantification of 14 antibacterial and antifungal agents (beta-lactams piperacillin, cefoperazone, and meropenem; beta-lactamase inhibitors tazobactam and sulbactam; antifungal agents fluconazole, caspofungin, posaconazole, and voriconazole; and daptomycin, vancomycin, teicoplanin, linezolid, and tigecycline) that can be used for patients with severe infection. This assay requires only 100µL of serum with rapid protein precipitation. Chromatographic analysis was performed using a Waters Acquity UPLC C8 column. Three stable isotope-labeled antibacterial agents and one analogue were used as internal standards. Calibration curves ranged from 0.1-100μg/mL, 0.1-50μg/mL, and 0.3-100μg/mL for different drugs, and all correlation coefficients were greater than 0.9085. Intra- and inter-day imprecision and inaccuracy values were below 15%. After validation, this new method was successfully employed for TDM in routine practice.

Suppressive effect of therapeutic antibiotic regimen on antipneumococcal Th1/Th17 responses in neonatal mice.

Antibiotics are commonly used in human neonates, but their impact on neonatal T cell immunity remains poorly understood. The aim of this study was to investigate the impact of the antibiotic piperacillin with the beta-lactamase inhibitor tazobactam on neonatal CD4+ and CD8+ T cell responses to Streptococcus pneumoniae. Splenic and lung cells were isolated from the neonatal mice receiving piperacillin and tazobactam or saline (sham) and cultured with S. pneumoniae to analyze T cell cytokine production by ELISA and flow cytometry. Antibiotic exposure to neonatal mice resulted in reduced numbers of CD4+/CD8+ T cells in the spleen and lungs compared to control mice. Upon in vitro stimulation with S. pneumoniae, splenocytes and lung cells from antibiotic-exposed mice produced lower levels of IFN-γ (Th1)/IL-17A (Th17) and IL-17A cytokines, respectively. Flow cytometric analysis revealed that S. pneumoniae-stimulated splenic CD4+ T cells from antibiotic-exposed mice expressed decreased levels of IFN-γ and IL-17A compared to control mice, whereas lung CD4+ T cells produced lower levels of IL-17A. However, no significant difference was observed for IL-4 (Th2) production. Neonatal mice exposure to piperacillin and tazobactam reduces the number of CD4+ and CD8+ T cells, and suppresses Th1 and Th17, but not Th2, responses to S. pneumoniae. Exposure of neonatal mice with a combination of piperacillin and tazobactam reduces CD4+/CD8+ T cells in the spleen and lungs. Antibiotic exposure suppresses neonatal Th1 and Th17, but not Th2, responses to Streptococcus pneumoniae. Our findings may have important implications for developing better therapeutic strategies in the neonatal intensive care unit.

A liquid chromatography-tandem mass spectrometry platform for the routine therapeutic drug monitoring of 14 antibiotics: Application to critically ill pediatric patients

BackgroundThe accurate measurement of plasma levels of antibiotics is crucial for the individualization of antimicrobial therapies based on PK/PD strategies. In this paper we describe a new rapid and simple LC–MS/MS platform for quantifying 14 antibiotics (amikacin, amoxicillin, ceftazidime, ciprofloxacin, colistin, daptomycin, gentamicin, linezolid, meropenem, piperacillin, teicoplanin, tigecycline, tobramycin and vancomycin) and a beta-lactamase inhibitor (tazobactam) starting from 50 μL plasma samples. MethodsAnalyses were performed on a Thermo Scientific™ Ultimate™ 3000 LC system (Thermo Fisher Scientific, Milan, Italy) coupled to a Thermo Scientific™ TSQ Quantiva™ Triple Quadrupole mass spectrometer. After fast protein precipitation protocols and addition of deuterated internal standards, samples were subjected to a fast HPLC gradient separation and the 15 drugs were quantified using multiple reaction monitoring of specific transitions over a wide range of concentrations. The suitability of the assay for TDM was tested on plasma samples derived from pediatric patients under treatment with one or more antibiotics. ResultsThe overall turnaround time of the assay was 20 min. The assay was validated following EMA guidelines for bioanalytical method validation and showed excellent accuracy (ranging from 85.3 and 112.7) and reproducibility (ranging from 1.3 to 9.7) as well as the absence of matrix effects (<15 %) for all the drugs tested. The lower limits of quantifications were between 0.1 and 2 mg/L. the recovery rate exceeded 85 % for all the drug tested. Stability was evaluated in different conditions thus allowing the setting up of reliable operative procedures. ConclusionsThis work provides a LC–MS/MS platform validated for clinical use for a rapid quantification of a broad spectrum of drugs having different chemical characteristics in a small volume of plasma and is suitable for real-time TDM-guided personalization of antimicrobial treatment in critically ill patients.

Molecular Dynamics Study of the Complex Formation of TEM-Type β-Lactamases with Substrates and Inhibitors

The complex formation of TEM-1 β-lactamase and its three mutant forms TEM-32, TEM-37, and TEM-39 with substrates cephalothin and CENTA and serine beta-lactamase inhibitors sulbactam, tazobactam, and clavulanic acid is studied using the methods of molecular dynamics. It is found that the stability of the complexes is caused by the electrostatic attraction between the deprotonated carboxyl group of the β-lactam ring of the substrate (inhibitor) and the positively charged amino groups of the lysine 234 and 73 residues, located in the active site of the enzymes. The formation of a hydrogen bond between this substrate group or its carbonyl oxygen with the hydroxyl group of the catalytic serine 70 residue and also between the negatively charged substituent groups and the positive charge region formed by the arginine 244 guanidine group and the asparagine 276 amino group is observed for some complexes. The binding energy of CENTA with TEM-1 β-lactamase is below the analogous binding energy of cephalothin, which is confirmed by the values of the Michaelis constants, determined experimentally. It is also found that the inhibitors bind to the mutant forms of β-lactamases related to the inhibitor-resistant phenotype, with higher affinity than TEM-1 β-lactamase.

Classical β-Lactamase Inhibitors Potentiate the Activity of Daptomycin against Methicillin-Resistant Staphylococcus aureus and Colistin against Acinetobacter baumannii.

ABSTRACTWe asked whether beta-lactamase inhibitors (BLIs) increased the activity of daptomycin (DAP) against methicillin-resistant Staphylococcus aureus (MRSA), the peptide antibiotic colistin (COL) against the emerging Gram-negative nosocomial pathogen Acinetobacter baumannii, and the human host defense peptide cathelicidin LL37 against either pathogen. DAP and LL37 kill curves were performed with or without BLIs against MRSA, vancomycin-intermediate S. aureus (VISA), and heterogeneous VISA (hVISA). COL and LL37 kill curves were performed against A. baumannii. Boron-dipyrromethene (BODIPY)-labeled DAP binding to MRSA grown with the BLI tazobactam (TAZ) was assessed microscopically. The combination of COL plus TAZ was studied in a murine model of A. baumannii pneumonia. TAZ alone lacked in vitro activity against MRSA or A. baumannii. The addition of TAZ to DAP resulted in a 2- to 5-log10 reduction in recoverable MRSA CFU at 24 h compared to the recoverable CFU with DAP alone. TAZ plus COL showed synergy by kill curves for 4 of 5 strains of A. baumannii tested. Growth with 20 mg/liter TAZ resulted in 2- to 2.5-fold increases in the intensity of BODIPY-DAP binding to MRSA and hVISA strains. TAZ significantly increased the killing of MRSA and A. baumannii by LL37 in vitro. TAZ increased the activity of COL in a murine model of A. baumannii pneumonia. Classical BLIs demonstrate synergy with peptide antibiotics. Since BLIs have scant antimicrobial activity on their own and are thus not expected to increase selective pressure toward antibiotic resistance, their use in combination with peptide antibiotics warrants further study.

Treatment of Skin and Soft Tissue Infections.

Bacterial infections of the skin range from mild pyodermas to life-threatening necrotizing infections. Pyodermas are most often due to Staphylococcus aureus or beta-hemolytic Streptococcus sp, whereas infections associated with skin ulcers of the extremities, infections following trauma or surgery, and histotoxic necrotizing infections may involve a large number of additional pathogens, including Enterobacteriaceae, Pseudomonas sp, enterococci, and anaerobes. Management of bacterial skin and soft-tissue infections includes appropriate surgical drainage or excision of infected tissue and antimicrobial therapy. The combination of piperacillin and the beta-lactamase inhibitor tazobactam is a newly released antimicrobial, which has excellent in vitro activity against the vast majority of pathogens involved in skin infections. Two multicenter studies recently evaluated the efficacy and safety of piperacillin/tazobactam in the therapy of skin and soft-tissue infections in hospitalized patients. Piperacillin/tazobactam was well tolerated and demonstrated high clinical efficacy for the treatment of these infections.

Current Challenges in Antimicrobial Chemotherapy

The use of the three classical beta-lactamase inhibitors (clavulanic acid, tazobactam and sulbactam) in combination with beta-lactam antibacterials is currently the most successful strategy to combat beta-lactamase-mediated resistance. However, these inhibitors are efficient in inactivating only class A beta-lactamases and the efficiency of the inhibitor/antibacterial combination can be compromised by several mechanisms, such as the production of naturally resistant class B or class D enzymes, the hyperproduction of AmpC or even the production of evolved inhibitor-resistant class A enzymes. Thus, there is an urgent need for the development of novel inhibitors. For serine active enzymes (classes A, C and D), derivatives of the beta-lactam ring such as 6-beta-halogenopenicillanates, beta-lactam sulfones, penems and oxapenems, monobactams or trinems seem to be potential starting points to design efficient molecules (such as AM-112 and LK-157). Moreover, a promising non-beta-lactam molecule, NXL-104, is now under clinical development. In contrast, an ideal inhibitor of metallo-beta-lactamases (class B) remains to be found, despite the huge number of potential molecules already described (biphenyl tetrazoles, cysteinyl peptides, mercaptocarboxylates, succinic acid derivatives, etc.). The search for such an inhibitor is complicated by the absence of a covalent intermediate in their catalytic mechanisms and the fact that beta-lactam derivatives often behave as substrates rather than as inhibitors. Currently, the most promising broad-spectrum inhibitors of class B enzymes are molecules presenting chelating groups (thiols, carboxylates, etc.) combined with an aromatic group. This review describes all the types of molecules already tested as potential beta-lactamase inhibitors and thus constitutes an update of the current status in beta-lactamase inhibitor discovery.

Three decades of beta-lactamase inhibitors.

Since the introduction of penicillin, beta-lactam antibiotics have been the antimicrobial agents of choice. Unfortunately, the efficacy of these life-saving antibiotics is significantly threatened by bacterial beta-lactamases. beta-Lactamases are now responsible for resistance to penicillins, extended-spectrum cephalosporins, monobactams, and carbapenems. In order to overcome beta-lactamase-mediated resistance, beta-lactamase inhibitors (clavulanate, sulbactam, and tazobactam) were introduced into clinical practice. These inhibitors greatly enhance the efficacy of their partner beta-lactams (amoxicillin, ampicillin, piperacillin, and ticarcillin) in the treatment of serious Enterobacteriaceae and penicillin-resistant staphylococcal infections. However, selective pressure from excess antibiotic use accelerated the emergence of resistance to beta-lactam-beta-lactamase inhibitor combinations. Furthermore, the prevalence of clinically relevant beta-lactamases from other classes that are resistant to inhibition is rapidly increasing. There is an urgent need for effective inhibitors that can restore the activity of beta-lactams. Here, we review the catalytic mechanisms of each beta-lactamase class. We then discuss approaches for circumventing beta-lactamase-mediated resistance, including properties and characteristics of mechanism-based inactivators. We next highlight the mechanisms of action and salient clinical and microbiological features of beta-lactamase inhibitors. We also emphasize their therapeutic applications. We close by focusing on novel compounds and the chemical features of these agents that may contribute to a "second generation" of inhibitors. The goal for the next 3 decades will be to design inhibitors that will be effective for more than a single class of beta-lactamases.

Detection of bacterial strains producing sulbactam‐ or tazobactam‐sensitive β‐lactamases by the use of disks containing the inhibitors alone instead of combining them with antibiotics

The author demonstrated that disks containing beta-lactamase inhibitors sulbactam or tazobactam combined with ampicillin (SAM) and piperacillin (TZP) are not suitable for performing the double-disk synergy test (DDST) with the aim of determining the sensitivity of beta-lactamases to these inhibitors. The presence of antibiotics (especially of piperacillin) is so disturbing that the results of testing are not specific. In contrast, the use of disks containing sulbactam or tazobactam alone yields very specific results. The author suggested to the firms producing sensi-disks that they make these commercially available to laboratory workers.

Activities of various β-lactams and β-lactam/β-lactamase inhibitor combinations against Acinetobacter baumannii and Acinetobacter DNA group 3 strains

Acinetobacter baumannii and Acinetobacter DNA group 3 are members of the so-called A. calcoaceticus-A. baumannii complex and are important nosocomial pathogens. Multiresistance in these organisms is increasingly frequent, and alternative treatment options are needed. The beta-lactamase inhibitors clavulanate, sulbactam and tazobactam have intrinsic activity against Acinetobacter strains. In the present study, broth microdilution was used to assess the in-vitro activities of currently available beta-lactam/beta-lactamase inhibitor combinations and sulbactam alone against 469 Acinetobacter isolates (A. baumannii, n=395; Acinetobacter DNA group 3, n=74) collected from various laboratories in Germany. Fixed concentrations and fixed ratios of beta-lactamase inhibitors were used. Sulbactam-containing combinations (susceptibility rates of 90.4-92.7% for A. baumannii and 97.3-100% for Acinetobacter DNA group 3) and sulbactam alone were superior to clavulanate- and tazobactam-containing combinations. The activity of sulbactam-containing combinations against members of the A. calcoaceticus-A. baumannii complex was conferred exclusively by the intrinsic activity of the beta-lactamase inhibitor and did not result from enhanced beta-lactam activity. Testing with the inhibitor added at a fixed ratio of inhibitor to beta-lactam appeared to give more reliable results than testing at a fixed concentration of the inhibitor. Resistance to carbapenems (0.3%) remains low in Germany.

The Penicillins

The penicillin family of antibiotics remains an important part of our antimicrobial armamentarium. In general, these agents have bactericidal activity, excellent distribution throughout the body, low toxicity, and efficacy against infections caused by susceptible bacteria. The initial introduction of aqueous penicillin G for treatment of streptococcal and staphylococcal infections was an important pharmacologic landmark. The emergence of penicillinase-producing Staphylococcus aureus prompted the development of the penicillinase-resistant penicillins (for example, methicillin, oxacillin, and nafcillin), in which an acyl side chain prevented disruption of the beta-lactamase ring. Subsequently, the aminopenicillins (ampicillin, amoxicillin, and bacampicillin) were developed because of the need for gram-negative antimicrobial activity. Their spectrum initially included Escherichia coli, Proteus mirabilis, Shigella, Salmonella, Listeria, Haemophilus, and Neisseria. The search for a penicillin with additional antimicrobial activity against the Enterobacteriaceae and Pseudomonas aeruginosa led to the development of the carboxypenicillins (carbenicillin and ticarcillin) and the ureidopenicillins (mezlocillin, azlocillin, and piperacillin). Finally, the combination of a beta-lactamase inhibitor (clavulanic acid, sulbactam, or tazobactam) and an aminopenicillin, ticarcillin, or piperacillin has further extended their antibacterial spectra by inhibiting certain beta-lactamases (non-group 1) of resistant bacteria. The development of an ideal penicillin that is rapidly bactericidal, nonsensitizing, nontoxic, bioavailable, and resistant to beta-lactamases and that has a high affinity for penicillin-binding proteins remains the goal.

Inhibitor resistant class A beta-lactamases

Beta-lactamase inhibitors (clavulanic acid, tazobactam, and sulbactam) greatly enhance the therapeutic efficacy of their partner antibiotics (amoxacillin, ampicillin, piperacillin, and ticarcillin) against common enteric and non-enteric organisms possessing class A beta-lactamases. Unfortunately, the number of class A enzymes being discovered that are resistant to these combinations is increasingly rapidly. The TEM and SHV class A beta-lactamases resistant to inhibitors have point mutations in critical amino acids important for catalysis. Compared to the wild type beta-lactamase, inhibitor resistant enzymes are inefficient at hydrolyzing benzylpenicillin, aminopenicillins, and cephalosporins. Nevertheless, hyper-production of these enzymes resulting from mutations in the promoter region can confer substantial levels of resistance. Understanding the microbiologic and kinetic properties of these inhibitor resistant class A beta-lactamases can lead to the design of more potent beta-lactam compounds as well as more effective inhibitors.

Inhibitor resistant class A beta-lactamases.

Beta-lactamase inhibitors (clavulanic acid, tazobactam, and sulbactam) greatly enhance the therapeutic efficacy of their partner antibiotics (amoxacillin, ampicillin, piperacillin, and ticarcillin) against common enteric and non-enteric organisms possessing class A beta-lactamases. Unfortunately, the number of class A enzymes being discovered that are resistant to these combinations is increasingly rapidly. The TEM and SHV class A beta-lactamases resistant to inhibitors have point mutations in critical amino acids important for catalysis. Compared to the wild type beta-lactamase, inhibitor resistant enzymes are inefficient at hydrolyzing benzylpenicillin, aminopenicillins, and cephalosporins. Nevertheless, hyper-production of these enzymes resulting from mutations in the promoter region can confer substantial levels of resistance. Understanding the microbiologic and kinetic properties of these inhibitor resistant class A beta-lactamases can lead to the design of more potent beta-lactam compounds as well as more effective inhibitors.

Therapy of soft tissue infections with piperacillin/tazobactam

The efficacy and safety of piperacillin in combination with the beta-lactamase inhibitor tazobactam were assessed in an open, multi-centre study of hospitalized patients with skin and soft tissue infections. A total of 136 patients from four countries were treated with piperacillin 4 g plus tazobactam 500 mg every 8 h. The mean duration of treatment was eight days. Clinical and bacteriological evaluations were performed one to three days and ten to 14 days post-therapy, respectively. Among 120 evaluable patients 93% were clinically cured or improved; 91% of bacteriologically evaluable patients were cured or improved. Eradication of the pathogens was observed in 85% and 95% of the evaluable patients at first and second follow-up. In 43% of the patients more than one pathogen was isolated from the wound. Persistence was commonest in patients in whom a complete surgical debridement could not be carried out. There were adverse events in 16 of the 136 patients, most commonly allergic reactions and diarrhoea. Six patients were withdrawn from the study because of side-effects.

Selection of variants of the TEM-1 beta-lactamase, encoded by a plasmid of clinical origin, with increased resistance to beta-lactamase inhibitors.

A TEM-1 derived variant beta-lactamase with increased resistance to beta-lactamase inhibitors was selected in vitro. The variant beta-lactamase was obtained from the parent strain Escherichia coli J62-2 containing resistance plasmid R1 which encodes the TEM-1 beta-lactamase. The variants were obtained by repeated subculture of E. coli J62-2 (R1) for five days in the presence of sub-inhibitory concentrations of amoxycillin plus clavulanic acid. Comparison of this variant beta-lactamase with a clinically isolated TEM derived beta-lactamase, with increased resistance to beta-lactamase inhibitors, suggests that they are the same enzyme. Both beta-lactamases have the same pI at 5.25 and have similar ID50 values for the beta-lactamase inhibitors clavulanic acid, sulbactam and tazobactam.

A new plasmidic cefotaximase from patients infected withSalmonella typhimurium

Salmonella typhimurium strains resistant to most beta-lactams, co-trimoxazole, tobramycin and gentamicin were isolated from patients in two hospitals in Buenos Aires, Argentina, beginning in August 1990. The patients were suffering from meningitis, septicaemia or enteritis. Therapy including ampicillin, ceftriaxone and gentamicin failed. The strains produced a plasmidic (pMVP-4) extended broad-spectrum beta-lactamase which is more active against cefotaxime than against ceftazidime (Vmax for cefotaxime 350 times higher than for ceftazidime). This cefotaximase demonstrates similarity to the previously described CTX-ase-M-1 from Escherichia coli, but it is distinctly different from CTX-ase-M-1 by its isoelectric point (7.9 for CTX-ase-M-2 in comparison with 8.9 for CTX-ase-M-1) as well as in its lower susceptibility to the beta-lactamase inhibitors sulbactam, clavulanic acid, tazobactam and BRL 42715. Thus, the beta-lactamase produced by S. typhimurium strains from Argentina appears to represent a new member (CTX-ase-M-2) of a novel group of plasmidic extended broad-spectrum beta-lactamases designated as cefotaximases.

Purification and characterization of an imipenem hydrolysing metallo-beta-lactamase from Bacteroides fragilis.

An imipenem resistant beta-lactamase producing strain of Bacteroides fragilis was isolated from a clinical specimen. The specific activity of the unpurified beta-lactamase was 5.5 U/mg protein. The beta-lactamase was purified 60-fold by Q Sepharose, Sephacryl S-300 and Mono Q column passages. The strain was able to inactivate imipenem and cefoxitin in broth cultures. The enzyme hydrolysed imipenem more rapidly than ampicillin, benzylpenicillin, cephalothin and cefoxitin. The activity of the enzyme was Zn2+ dependent and was completely inhibited by EDTA. The inhibition was reversed by ZnSO4. Preincubation with the common beta-lactamase inhibitors clavulanic acid, sulbactam and tazobactam did not reduce the enzyme activity. The molecular weight was determined by sodium dodecyl sulfate gradient gel electrophoresis to be 31,000 Daltons and the isoelectric point was 4.5.

Comparison of the inoculum effect of cefoxitin and other cephalosporins and of beta-lactamase inhibitors and their penicillin-derived components on the Bacteroides fragilis group

We compared the inoculum effects for 109 recent clinical isolates of the Bacteroides fragilis group of cefoxitin, cefotetan, ceftizoxime, ceftriaxone, and three beta-lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) and their penicillin-derived components. Bactericidal activity was assayed and morphologic changes were monitored for selected strains exhibiting a large inoculum effect. Ceftizoxime demonstrated the largest inoculum effect, followed by cefotetan and ceftriaxone. The large inoculum effect of ceftizoxime and ceftriaxone was correlated with filamentous transformation at the high inoculum (10(8) CFU/ml) and lack of bactericidal activity suggesting drug destruction or inactivation. Cefotetan was bactericidal for B. fragilis isolates but not for other members of the B. fragilis group. Cefoxitin showed the least inoculum effect and was consistently bactericidal at high (10(8) CFU/ml), standard (10(6) CFU/ml), and low (10(4) CFU/ml) inocula, followed by ampicillin-sulbactam. Piperacillin-tazobactam and ticarcillin-clavulanic acid showed an intermediate inoculum effect. The degree of inoculum effect observed generally correlated with bactericidal activity at all inocula.

Comparison of the inoculum effects of members of the family Enterobacteriaceae on cefoxitin and other cephalosporins, beta-lactamase inhibitor combinations, and the penicillin-derived components of these combinations

We compared the inoculum effects of 105 recent clinical isolates of the family Enterobacteriaceae on cefoxitin, other cephalosporins, aztreonam, and three beta-lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) and their penicillin-derived components. Piperacillin and aztreonam showed the largest inoculum effect, and cefoxitin showed the smallest. The other cephalosporins tested (cefotetan, ceftizoxime, and ceftriaxone) showed an intermediate inoculum effect. In general, the inoculum effect was of greater magnitude for the penicillin and beta-lactamase inhibitor combinations than for the cephalosporins tested. Bactericidal activity was assayed and morphologic changes were monitored for selected strains exhibiting a large inoculum effect. MICs correlated with bactericidal activity at an inoculum level of 10(5) CFU/ml, while activity at 10(8) CFU/ml was variable. Cefoxitin demonstrated the least filamentous transformation and the most rapid bactericidal activity. Aztreonam showed the most marked filamentous transformation and was no longer bactericidal at 10(8) CFU/ml. The beta-lactamase inhibitor combinations showed variable bactericidal activity, and regrowth occurred with a number of strains with all three agents tested.

Beta-lactam-fosfomycin antagonism involving modification of penicillin-binding protein 3 in Pseudomonas aeruginosa

Antagonism between fosfomycin and antipseudomonal penicillins, cefotaxime, and ceftriaxone was observed in Pseudomonas aeruginosa RYC212. Fosfomycin, a non-beta-lactam antibiotic that acts on bacterial cell wall synthesis, decreased the expression of penicillin-binding protein 3 and induced beta-lactamase. The antagonistic effect was reduced in the presence of high concentrations of the beta-lactamase inhibitor tazobactam or in fosfomycin-resistant mutants. We suggest that products resulting from fosfomycin cell wall damage could interact with a system that regulates penicillin-binding protein and beta-lactamase production.