Skip to main content

Relebactam restores susceptibility of resistant Pseudomonas aeruginosa and Enterobacterales and enhances imipenem activity against chromosomal AmpC-producing species: analysis of global SMART 2018–2020

Abstract

Background

Carbapenem-resistant bacteria are an increasing problem in clinical practice; thus, it is important to identify β-lactamase inhibitors (e.g., relebactam) that can restore carbapenem susceptibility. We report analyses of relebactam enhancement of imipenem activity against both imipenem-nonsusceptible (NS) and imipenem-susceptible (S) Pseudomonas aeruginosa and Enterobacterales. Gram-negative bacterial isolates were collected for the ongoing Study for Monitoring Antimicrobial Resistance Trends global surveillance program. Clinical and Laboratory Standards Institute–defined broth microdilution minimum inhibitory concentrations (MIC) were used to determine the imipenem and imipenem/relebactam antibacterial susceptibilities of P. aeruginosa and Enterobacterales isolates.

Results

Between 2018 and 2020, 36.2% of P. aeruginosa (N = 23,073) and 8.2% of Enterobacterales (N = 91,769) isolates were imipenem-NS. Relebactam restored imipenem susceptibility in 64.1% and 49.4% of imipenem-NS P. aeruginosa and Enterobacterales isolates, respectively. Restoration of susceptibility was largely observed among K. pneumoniae carbapenemase-producing Enterobacterales and carbapenemase-negative P. aeruginosa. Relebactam also caused a lowering of imipenem MIC among imipenem-S P. aeruginosa and Enterobacterales isolates from chromosomal Ambler class C β-lactamase (AmpC)–producing species. For both imipenem-NS and imipenem-S P. aeruginosa isolates, relebactam reduced the imipenem MIC mode from 16 μg/mL to 1 μg/mL and from 2 μg/mL to 0.5 μg/mL, respectively, compared with imipenem alone.

Conclusions

Relebactam restored imipenem susceptibility among nonsusceptible isolates of P. aeruginosa and Enterobacterales and enhanced imipenem susceptibility among susceptible isolates of P. aeruginosa and isolates from Enterobacterales species that can produce chromosomal AmpC. The reduced imipenem modal MIC values with relebactam may result in a higher probability of target attainment in patients.

Peer Review reports

Introduction

Carbapenems, members of the class of broad-spectrum antibacterial agents known as β-lactams, are an option for multidrug-resistant infections that may fail initial lines of therapy [1]. Nonsusceptibility to carbapenem treatment is frequently due to synergistic resistance mechanisms present within certain pathogenic strains (e.g., concurrent porin loss, modification of penicillin-binding proteins, and/or expression of β-lactamases [expanded spectrum or Ambler class C (AmpC), including Pseudomonas-derived cephalosporinase (PDC)]) [2, 3]. AmpC can either be encoded by chromosomal genes, inducible upon exposure to certain β-lactam antibacterial agents, such as imipenem [4], or plasmid-acquired and generally constitutively expressed [5]. Chromosomal AmpC is a particularly important mechanism of resistance for numerous β-lactam antibacterial agents in the treatment of P. aeruginosa [6]. In addition, among Enterobacterales, certain species such as Citrobacter freundii, Enterobacter cloacae, Klebsiella aerogenes, and Serratia marcescens encode a chromosomal AmpC enzyme whose expression can be de-repressed, either by genetic mutation or the presence of an inducing β-lactam antibacterial, such as imipenem [6]. Some carbapenem-resistant Enterobacterales isolates do not produce a carbapenemase, and resistance is due to the presence of an extended spectrum β-lactamase or AmpC enzyme in combination with loss of expression of outer membrane porins. In addition, certain physiologic conditions among critically ill patients, such as augmented renal clearance (ARC), may lead to underdosing, which can contribute to inadequate response to therapy [7, 8]. Some β-lactams are particularly susceptible to subtherapeutic treatment exposures that may result from ARC, especially among isolates with minimum inhibitory concentrations (MIC) at the higher end of the susceptibility range, limiting their bactericidal activity [7, 8]. Development of suitable β-lactam/β-lactamase inhibitor combinations, such as imipenem/cilastatin/relebactam (IMI/REL), capable of overcoming loss of carbapenem susceptibility and limiting potential for underexposure, is important because of the ongoing global threat of multidrug-resistant bacteria and the potential for inadequate dosing in critically ill patients [7,8,9,10,11,12,13].

Relebactam is an inhibitor of Ambler class A and class C (e.g., AmpC) β-lactamases that, when combined with imipenem, restores imipenem activity against nonsusceptible isolates and enhances imipenem activity, specifically against susceptible P. aeruginosa isolates [2]. In a fixed-dose combination with imipenem/cilastatin, relebactam was approved in the United States and European Union for hospital-acquired pneumonia and ventilator-associated pneumonia, bacteraemia associated with hospital-acquired pneumonia/ventilator-associated pneumonia (European Union only), and infections due to aerobic gram-negative organisms in adults with limited treatment options (e.g., complicated urinary tract infections [cUTI] and complicated intra-abdominal infections [cIAI]) [14, 15].

Previous analysis of in vitro activity in imipenem-nonsusceptible (NS) isolates indicated that relebactam lowers MICs through inhibition of β-lactamase activity [2, 16]. Here, we expand upon previous reports of relebactam potentiation of imipenem activity against both imipenem-NS and imipenem-susceptible (S) P. aeruginosa surveillance isolates and extend this analysis to both imipenem-NS and imipenem-S Enterobacterales isolates from the ongoing Study for Monitoring Antimicrobial Resistance Trends (SMART) global surveillance program [16, 17]. The SMART program was initiated in 2002 and includes collection and assessment of clinical isolates by hospital laboratories for monitoring antibacterial susceptibility profiles of gram-negative bacteria [17].

Results

Between 2018 and 2020, 23,073 P. aeruginosa isolates and 91,769 Enterobacterales isolates were collected from patients at sites participating in the SMART program. For P. aeruginosa, 36.2% (n = 8356) of isolates were classified as imipenem-NS according to the Clinical and Laboratory Standards Institute (CLSI) breakpoint (MIC > 2 μg/mL). For Enterobacterales, 8.2% (n = 7493) of isolates were classified as imipenem-NS according to the CLSI breakpoint (MIC > 1 μg/mL).

Among P. aeruginosa isolates (N = 23,073), the presence of relebactam increased imipenem susceptibility from 63.8% to 87.0% (Fig. 1A) and reduced the MIC50/90 from 2/32 to 0.5/4 µg/mL; the mode MIC was reduced from 2 to 0.5 µg/mL (Table 1). The addition of relebactam restored imipenem susceptibility to 5353 (64.1%) of 8356 imipenem-NS P. aeruginosa isolates (Fig. 1B), reduced the MIC50 from 16 to 2 µg/mL and the mode MIC from 16 to 1 µg/mL (Table 1). Among molecularly characterized isolates, carbapenemases were rarely identified (0.2%) in imipenem-NS isolates for which relebactam restored susceptibility, and metallo-β-lactamases (MBL) were the most common carbapenemase (37.3%) in imipenem-NS isolates for which relebactam did not restore imipenem susceptibility (see Additional file 1). In addition, relebactam enhanced imipenem susceptibility among isolates of P. aeruginosa classified as imipenem-S (n = 14,717) by causing a shift toward lower MIC values (Fig. 1C), with the MIC50/90 decreasing from 1/2 µg/mL for imipenem alone to 0.5/0.5 µg/mL for imipenem/relebactam; the mode MIC was reduced from 2 µg/mL for imipenem to 0.5 µg/mL for imipenem/relebactam (Table 1).

Fig. 1
figure 1

Relebactam restores/enhances the activity of imipenem in Pseudomonas aeruginosa isolates. A all (N = 23,073). B imipenem-NS (N = 8356). C imipenem-S (N = 14,717). Percentage represents n/N × 100%, where n was the number of isolates meeting the MIC threshold and N was the total number of isolates based on the CLSI 2021 clinical breakpoints for imipenem/relebactam (MIC ≤ 2 μg/mL for susceptibility) and subsequently categorized as either S (MIC ≤ 2 μg/mL) or NS (MIC > 2 μg/mL) [18]. The dashed line indicates the CLSI 2021 imipenem/relebactam susceptibility breakpoints. The arrows indicate mode MIC values. CLSI, Clinical and Laboratory Standards Institute; MIC, minimum inhibitory concentration; NS, nonsusceptible; S, susceptible

Table 1 MIC50/90 and mode MIC values for imipenem-S, imipenem-NS, and total Pseudomonas aeruginosa and Enterobacterales.a

Among all Enterobacterales isolates (N = 91,769), the addition of relebactam increased imipenem susceptibility from 91.8% to 95.8% (see Additional file 2A) and reduced the MIC50/90 from 0.25/1 to ≤ 0.12/0.5 µg/mL (Table 1). For the 7493 isolates of Enterobacterales classified as imipenem-NS, the addition of relebactam restored imipenem susceptibility in 3704 (49.4%) isolates (see Additional file 2B) and reduced the MIC50 from > 8 to 2 µg/mL (Table 1). Among molecularly characterized isolates for which relebactam restored imipenem susceptibility, a majority (52.4%) encoded K. pneumoniae carbapenemases (KPCs), whereas MBLs (53.6%) and oxacillinase (OXA)-48 family β-lactamases (44.0%) were common among imipenem-NS isolates for which relebactam did not restore imipenem susceptibility. In addition, relebactam enhanced imipenem susceptibility among isolates of Enterobacterales classified as imipenem-S (N = 84,276) by inducing a shift toward lower MIC values (see Additional file 2C), with the MIC50 being reduced from 0.25 to ≤ 0.12 µg/mL (Table 1).

Because AmpC-producing Enterobacterales species have reduced susceptibility to imipenem [2, 19] and imipenem is a potent inducer of AmpC expression [4], we further analysed Enterobacterales species as separate subgroups based upon their capacity for chromosomal AmpC production. The chromosomal AmpC producers were E. cloacae, S. marcescens, K. aerogenes, and C. freundii; the nonproducers were E. coli, K. pneumoniae, K. oxytoca, and C. koseri. Among all isolates from chromosomal AmpC–producing species (n = 13,003), the addition of relebactam increased imipenem susceptibility from 87.4% to 95.4% (Fig. 2A) and reduced the MIC50/90 from 0.5/2 to 0.25/1 µg/mL; in addition, the mode MIC was reduced from 0.5 to 0.25 µg/mL (Table 1). Among imipenem-NS isolates from chromosomal AmpC–producing species (n = 1634), the addition of relebactam restored susceptibility to 64.2% of isolates (Fig. 2B), reduced the MIC50 from 2 to 1 µg/mL, and the mode MIC from 2 to 0.25 µg/mL (Table 1). Molecular characterization of imipenem-NS isolates for which relebactam restored imipenem susceptibility found that carbapenemases were present in 16.7% of isolates, with KPC as the most common carbapenemase, present in 13.7% of isolates (see Additional file 1). MBLs were present in 59.4% of imipenem-NS isolates for which relebactam did not restore imipenem susceptibility (see Additional file 1). With regards to imipenem-S isolates from chromosomal AmpC–producing species (n = 11,369), the addition of relebactam caused a shift towards reduced MICs (Fig. 2C) and reduced the MIC50/90 from 0.5/1 to 0.25/0.5 µg/mL; in addition, the mode MIC was reduced from 0.5 to 0.25 µg/mL (Table 1).

Fig. 2
figure 2

Relebactam restores the activity of imipenem in chromosomal AmpC–producing Enterobacterales isolates. A all (N = 13,003). B imipenem-NS (N = 1634). C imipenem-S (N = 11,369). Percentage represents n/N × 100%, where n was the number of isolates meeting the MIC threshold and N was the total number of isolates based on the CLSI 2021 clinical breakpoints for imipenem and imipenem/relebactam (both MIC ≤ 1 μg/mL for susceptibility) and subsequently categorized as either S (MIC ≤ 1 μg/mL) or NS (MIC > 1 μg/mL) [18]. The dashed line indicates the CLSI 2021 imipenem and imipenem/relebactam susceptibility breakpoints. The arrows indicate mode MIC values. Enterobacterales chromosomal AmpCproducing species included Enterobacter cloacae, Serratia marcescens, Klebsiella aerogenes, and Citrobacter freundii. AmpC, Ambler class C β-lactamase; CLSI, Clinical and Laboratory Standards Institute; MIC, minimum inhibitory concentration; NS, nonsusceptible; S, susceptible

Among all Enterobacterales isolates from chromosomal AmpC–nonproducing species (N = 78,766) (Fig. 3A), the addition of relebactam increased imipenem susceptibility from 92.6% to 95.9% and reduced the MIC50/90 from 0.25/1 to ≤ 0.12/0.5 µg/mL (Table 1). Among imipenem-NS isolates from chromosomal AmpC–nonproducing species (n = 5859), the addition of relebactam restored susceptibility to 45.3% of isolates (Fig. 3B) and reduced the MIC50 from > 8 to 2 µg/mL (Table 1). Molecular characterization of these isolates found that KPC was present in 68.1% of isolates for which relebactam restored imipenem susceptibility; among isolates for which relebactam did not restore imipenem susceptibility, 36.1% encoded an OXA-48 family β-lactamase, 40.0% encoded an MBL, and 11.8% encoded both an OXA-48 family β-lactamase and an MBL (see Additional file 1). Among imipenem-S isolates from chromosomal AmpC–nonproducing species (n = 72,907), the addition of relebactam resulted in a small downward shift in MICs (Fig. 3C) and reduced the MIC90 from 0.5 to 0.25 µg/mL (Table 1).

Fig. 3
figure 3

Relebactam enhances the activity of imipenem among chromosomal AmpC–nonproducing Enterobacterales isolates. A all (N = 78,766). B imipenem-NS (N = 5859). C imipenem-S (N = 72,907). Percentage represents n/N × 100%, where n was the number of isolates meeting the MIC threshold and N was the total number of isolates based on the CLSI 2021 clinical breakpoints for imipenem and imipenem/relebactam (both MIC ≤ 1 μg/mL for susceptibility) and subsequently categorized as either S (MIC ≤ 1 μg/mL) or NS (MIC > 1 μg/mL) [18]. The dashed line indicates the CLSI 2021 imipenem and imipenem/relebactam susceptibility breakpoints. The arrows indicate mode MIC values. Enterobacterales chromosomal AmpCnonproducing species included Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca, and Citrobacter koseri. AmpC, Ambler class C β-lactamase; CLSI, Clinical and Laboratory Standards Institute; MIC, minimum inhibitory concentration; NS, nonsusceptible; S, susceptible

The MIC data were also evaluated using imipenem European Committee on Antimicrobial Susceptibility Testing (EUCAST) susceptibility breakpoints of for Enterobacterales and 4 µg/mL for P. aeruginosa and imipenem/relebactam susceptibility breakpoints of 2 µg/mL for both Enterobacterales and P. aeruginosa (Table 1). The summary statistics using either CLSI or EUCAST breakpoints were largely similar for P. aeruginosa and Enterobacterales; they were either identical or within a single MIC dilution regardless of which interpretive criteria were applied. One notable difference was that for imipenem nonsusceptible isolates from chromosomal AmpC-producing Enterobacterales species, the imipenem mode MIC increased from 2 µg/mL using CLSI criteria to > 8 µg/mL using EUCAST criteria, as those isolates with MICs of 2 µg/mL were categorized as susceptible. Although there was no longer a reduction in the imipenem mode MIC in the presence of relebactam for these isolates, the MIC50 was still reduced by at least two dilutions, from > 8 to 4 µg/mL.

We evaluated the imipenem/relebactam susceptibility of imipenem-nonsusceptible isolates by region because of the geographic variation in the prevalence of carbapenemase enzymes among Enterobacterales inhibited by relebactam (i.e., KPC) and those not inhibited by relebactam (i.e., MBLs and OXA-48 family β-lactamases)[20], (Additional file 3). Relebactam restored imipenem susceptibility to > 70% of imipenem-nonsusceptible Enterobacterales isolated from Latin America, North America, or the South Pacific, where isolates frequently encode KPC or are carbapenemase negative. In Asia and Europe, where OXA-48 family β-lactamases and MBLs, respectively, are more common, relebactam restored imipenem susceptibility to 42.0% and 45.5%, respectively, of imipenem-NS Enterobacterales isolates. In addition, relebactam restored imipenem susceptibility to 63.7% to %-75.1% of imipenem-NS P. aeruginosa isolates from Asia, Europe, the Middle East, and North America, as well as 91.9% of isolates from the South Pacific. The rate of imipenem/relebactam susceptibility among imipenem-NS P. aeruginosa isolates was lower for isolates from Africa (48.3%) and Latin America (52.2%).

Discussion

The present study expanded analysis of previous findings demonstrating that relebactam restores imipenem activity in imipenem-NS Enterobacterales and P. aeruginosa isolates and enhances imipenem activity in imipenem-S P. aeruginosa isolates [16]. In addition, this study extended the analysis of relebactam with imipenem among Enterobacterales species and evaluated Enterobacterales species as a function of their capacity to produce chromosomally encoded AmpC.

Among imipenem-NS isolates, relebactam restored susceptibility in chromosomal AmpC–nonproducing species (e.g., K. pneumoniae), largely by inhibition of KPC, and in chromosomal AmpC–producing species (e.g., E. cloacae), presumably through inhibition of the chromosomal AmpC enzyme. Relebactam restored imipenem susceptibility to approximately two-thirds and one-half of chromosomal AmpC–producing and AmpC–nonproducing Enterobacterales species, respectively, demonstrating that relebactam-mediated inhibition of β-lactamase activity can prevent loss of carbapenem susceptibility. Notable exceptions to the decreases in MIC values observed with imipenem/relebactam occurred in isolates encoding certain β-lactamases (i.e., MBL and OXA). These isolates were minimally affected or unaffected by relebactam, which is consistent with lack of inhibition of class B and class D β-lactamases [14]. Collectively, these observations support previous findings that relebactam at a concentration of 4 µg/mL lowered imipenem MIC values [2, 16].

The mechanism of action for relebactam is inhibition of Class A or Class C β-lactamases to facilitate restoration or enhancement of imipenem susceptibility in gram-negative bacteria (Fig. 4; Additional file 4). Imipenem enters the periplasm through outer membrane porins [21]. Figure 4A depicts the effects upon addition of relebactam to imipenem among imipenem-NS P. aeruginosa and Enterobacterales isolates. In the absence of acquired carbapenemases (e.g., KPC, MBL, etc.), imipenem nonsusceptibility among P. aeruginosa and Enterobacterales is due to two factors: 1) loss of the imipenem entry porins (e.g., OprD, OmpK36, OmpF), which reduces entry of imipenem into the periplasm and 2) induced expression of the chromosomally encoded AmpC β-lactamase, which, although an inefficient carbapenemase, can degrade this reduced concentration of imipenem. Imipenem is a potent inducer of AmpC β-lactamases; therefore, whenever imipenem is present in a patient or an in vitro assay, AmpC will be hyperproduced [14, 21]. Relebactam likely restores imipenem susceptibility to these isolates by inhibiting AmpC, thereby allowing imipenem, which has entered the cell through nonspecific porins, to reach the target penicillin-binding proteins and exert its antibacterial effect, as observed for P. aeruginosa [16]. Among surveillance and genetically modified isolates, it is important to note that neither imipenem nor relebactam are substrates of efflux pumps [16, 22].

Fig. 4
figure 4

Effect of relebactam on susceptibility of Pseudomonas aeruginosa and Enterobacterales species to imipenem. A Relebactam restores susceptibility to imipenem in imipenem-NS Pseudomonas aeruginosa and Enterobacterales species. B Relebactam decreases imipenem MIC values in imipenem-S species. C These effects result in increased free drug pharmacokinetic/pharmacodynamic target (%ƒT > MIC) attainment for imipenem. %ƒT, percentage of time of free drug; AmpC, Ambler class C β-lactamase; AUC, area under the concentration–time curve; MIC, minimum inhibitory concentration; NS, nonsusceptible; S, susceptible

In contrast to imipenem-NS isolates, imipenem access to the periplasmic space through outer membrane porins (e.g., OprD, OmpK36, OmpF) is efficient in imipenem-S isolates, and the greater concentration of imipenem in the periplasm results in an antibacterial effect, even in the presence of induced AmpC (Fig. 4B) [21]. Chromosomal AmpC expression leads to an increase in imipenem MIC values without resulting in loss of susceptibility. In the presence of relebactam, the slow hydrolysis of imipenem by chromosomal AmpC is impeded and the MIC values of imipenem are thereby lowered, enhancing imipenem susceptibility [16].

The clinical implications of this lowered imipenem/relebactam MIC, compared with imipenem alone for both P. aeruginosa and the Enterobacterales are informative for an evaluation of adequacy of dose from a pharmacokinetic (PK)/pharmacodynamic (PD) perspective. The importance of achieving established PD targets with standard dosing regimens of older β-lactams has been heightened in recent years, given reports of underdosing of these β-lactam agents, particularly among critically ill patients with ARC [7, 8]. Concerns regarding underdosing have led to multiple changes that have had implications in routine clinical practice, such as lowering of MIC susceptibility breakpoints as a method to ensure higher doses are administered [23] and recommendations to administer prolonged or continuous infusions of β-lactams to increase the probability that adequate PD exposures are achieved [24, 25]. With this context in mind, the established PK/PD target for imipenem, and all carbapenems, is the percentage of time the free drug concentration remains above the MIC of the infecting organism (%fT > MIC), with maximum bactericidal effect achieved at an %ƒT > MIC of 30%–40% [26,27,28,29]. For the β-lactamase inhibitor relebactam, a ratio of the area under the unbound concentration–time curve to the MIC (fAUC/MIC) of 7.5 was associated with two-log kill in preclinical models [28,29,30,31]. High (> 90%) joint probability of target attainment for both imipenem and relebactam has been described for the 1.25-g IMI/REL dose (500 mg imipenem/500 mg cilastatin/250 mg relebactam) at an imipenem/relebactam MIC breakpoint of ≤ 2 µg/mL, which covers P. aeruginosa and Enterobacterales [24, 25]. Because the MIC is in the denominator of each PK/PD target (e.g., fT > MIC and fAUC/MIC), the addition of relebactam as a strategy to lower the imipenem MIC is expected to result in higher PD exposures (Fig. 4C) and serves as an alternative approach to extending the infusion to attain higher PD exposures. Although purely speculative, higher exposures achieved upon the addition of relebactam to imipenem may have contributed to the efficacy observed in the RESTORE-IMI 1 and 2 studies [10, 12, 14, 32, 33]. Of particular importance are the similar 28-day all-cause mortality and favorable clinical and microbiologic response rates among patients with normal renal function compared with those with ARC (creatinine clearance ≥ 150 mL/min) among participants with hospital-acquired/ventilator-associated bacterial pneumonia and ARC from the RESTORE-IMI 2 study [13].

In this study, molecularly characterized imipenem-nonsusceptible Enterobacterales isolates that remained nonsusceptible in the presence of relebactam frequently encoded an OXA-48 family β-lactamase, illustrating the lack of activity of relebactam against these enzymes. Among the few isolates in which relebactam restored susceptibility, the vast majority (84%) had MICs of , interpreted as intermediate susceptibility by CLSI and susceptible by EUCAST, which was attributable to the weak carbapenemase activity of OXA-48 family β-lactamases [34]. Imipenem-susceptible isolates were not characterized in this study; however, Enterobacterales isolates encoding OXA-48 family β-lactamases are frequently susceptible to meropenem [35], suggesting these enzymes are likely present in imipenem-susceptible isolates as well. Attributing the intermediate imipenem susceptibility phenotype to the presence of an OXA-48 family β-lactamase in a particular isolate is challenging; MIC values frequently vary by a single dilution in testing, and the presence of additional β-lactamases and resistance mechanisms (e.g., outer membrane porin loss) may be contributing factors. Overall, the results of this study reinforce prior findings that relebactam is an inhibitor class A/C β-lactamase and can restore imipenem susceptibility to isolates encoding these enzymes; correspondingly, the lack of relebactam activity toward class B/D β-lactamases is illustrated by the lack of meaningful restoration of imipenem susceptibility to isolates encoding these enzymes. From a clinical perspective, PK/PD data indicate that IMI/REL achieves high probability of target attainment for isolates with imipenem/relebactam MICs ≤ 2 µg/mL (i.e., the EUCAST susceptible, standard-dosing regimen breakpoint) [33], and limited clinical data indicate favorable clinical and microbiologic outcomes in a small number of trial participants (n = 3) with imipenem/relebactam-susceptible isolates encoding OXA-48 family β-lactamases (unpublished data) [12].

A limitation of the study was the range of MICs assessed. Wider ranges may have allowed detection of larger modal shifts (e.g., among imipenem-NS Enterobacterales). The relationship between bacterial susceptibility and antibacterial agent is complex; therefore, MIC may not be the best indicator of effectiveness of a particular antibacterial agent [36]. In addition, we did not directly measure AmpC production, but it is well recognized that this is one of the primary mechanisms of resistance of P. aeruginosa and certain Enterobacterales species [19, 24, 37] and that relebactam inhibition of chromosomally encoded AmpC enzymes is responsible for reduction in imipenem MIC values when acquired β-lactamases are not present [16].

The results of the present study indicated that encoded MBL and OXA genes contributed to certain Enterobacterales isolates remaining imipenem-NS after the addition of relebactam; however, the possibility remains that unidentified resistance mechanisms within these isolates contributed to their phenotype. The characterisation needed to confirm the presence of other resistance pathways was beyond the scope of the present study. Furthermore, although imipenem/relebactam circumvents certain resistance mechanisms in vitro with the associated decrease in MIC described here, other patient-specific factors may impact effectiveness to a greater extent.

Conclusions

Relebactam inhibits the ability of AmpC and KPC β-lactamases to hydrolyse imipenem in vitro, thereby restoring imipenem susceptibility among nonsusceptible isolates and enhancing imipenem susceptibility among susceptible isolates of P. aeruginosa and Enterobacterales. The reduction of imipenem modal MIC values with relebactam may result in a higher probability of target attainment in patients.

Materials and methods

Between 2018 and 2020, 243 unique participating sites collected up to 250 consecutive isolates each of aerobic gram-negative bacteria per year for the SMART program. Participating sites were located in 219 cities across 60 countries. The following number of isolates were collected from adult patients (≥ 18 years of age) at each site, each year hospitalized with: cUTIs (n = 50), cIAIs (n = 50), lower respiratory tract infections (n = 100), or bloodstream infections (n = 50). One isolate per species per patient per year was included. After collection, isolates were submitted to a central laboratory (International Health Management Associates, Inc [IHMA], Schaumburg, Illinois, USA) for analysis. All methods were carried out in accordance with the ethical principles Declaration of Helsinki and all relevant guidelines and regulations. Ethical approval and informed consent were not required because all isolates received into the study followed multiple subcultures and were completely de-identified. The secondary research use of de-identified isolates is considered exempt research according to the Regulations for the Protection of Human Subjects in Research of the U.S. Department of Health and Human Services, Office for Human Research Protections (45 CFR 46).

Confirmation of P. aeruginosa and Enterobacterales isolate identity was performed by IHMA using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (Bruker Daltonics, Billerica, Massachusetts, USA). Based on current IMI/REL indications, Enterobacterales species included in this analysis were Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Serratia marcescens, Klebsiella aerogenes, Klebsiella oxytoca, and Citrobacter freundii. In addition, Citrobacter koseri was also included because it is a carbapenem-resistant species that increasingly has been seen in hospital settings [38]. Isolates of Enterobacter cloacae, Serratia marcescens, Klebsiella aerogenes, and Citrobacter freundii were categorized as potential AmpC-producing species based on the presence of an AmpC-encoding gene and the potential for derepression due to genetic mutation or the presence of an inducing β-lactam antibacterial agent, such as imipenem [6].

Antibacterial susceptibility testing was performed at IHMA using CLSI standard broth microdilution methods [39]. The appropriate American Type Culture Collection control strains were used each day as quality-control measures in accordance with CLSI guidelines. Isolates were tested for susceptibility to both imipenem alone and imipenem/relebactam. Per CLSI recommendations, imipenem was diluted according to a two-fold gradient and tested in combination with a fixed concentration (4 μg/mL) of relebactam [18]. For P. aeruginosa, the CLSI breakpoints for imipenem and imipenem/relebactam susceptibility were both  [18]. The CLSI breakpoints for imipenem and imipenem/relebactam susceptibility were both ≤ 1 μg/mL for Enterobacterales [18]. Isolates with MIC values that exceeded these CLSI breakpoints were deemed nonsusceptible. The range of imipenem and imipenem/relebactam MICs tested for P. aeruginosa was ≤ 0.12 to > 32 µg/mL and for Enterobacterales was ≤ 0.12 to > 8 µg/mL.

In this study, isolates of P. aeruginosa and Enterobacterales classified as nonsusceptible to imipenem that were characterized molecularly for gene-encoded β-lactamases using previously described multiplex polymerase chain reaction assays and full-gene DNA sequencing techniques [40, 41] were evaluated. Over the current study period (2018–2020), 75% of imipenem-NS isolates from the species evaluated were analysed. Screening included assessment for gene-encoded MBLs, including imipenemase, Verona integron-encoded metallo-β-lactamase, New Delhi metallo-β-lactamase, and São Paulo MBL; serine β-lactamases (KPC; OXA) and chromosomally encoded PDC. The number of imipenem-nonsusceptible isolates molecularly characterized from the evaluated species is provided in Additional file 1. Isolates in which no carbapenemase-encoding genes were identified were characterized as carbapenemase negative. Gene-flanking primers were used to amplify and sequence (Sanger) all detected genes encoding carbapenemases and PDC for all Enterobacterales isolates in the study and for P. aeruginosa isolates from 2018 to 2019. P. aeruginosa isolates collected in 2020 that met the screening criteria were characterized by short-read whole-genome sequencing (Illumina HiSeq 2 × 150 base-pair reads) to a targeted coverage depth of 100 × [42] and analyzed using the CLC Genomics Workbench (Qiagen, Germantown, Maryland, USA). The ResFinder database was used to detect β-lactamase genes [43].

Availability of data and materials

Datasets used and analyzed for this study are available from the corresponding author upon reasonable request.

References

  1. Papp-Wallace KM, Endimiani A, Taracila MA, Bonomo RA. Carbapenems: past, present, and future. Antimicrob Agents Chemother. 2011;55(11):4943–60. https://doi.org/10.1128/AAC.00296-11.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Livermore DM, Warner M, Mushtaq S. Activity of MK-7655 combined with imipenem against Enterobacteriaceae and Pseudomonas aeruginosa. J Antimicrob Chemother. 2013;68(10):2286–90. https://doi.org/10.1093/jac/dkt178.

    Article  CAS  PubMed  Google Scholar 

  3. Rodríguez-Martínez JM, Poirel L, Nordmann P. Extended-spectrum cephalosporinases in Pseudomonas aeruginosa. Antimicrob Agents Chemother. 2009;53(5):1766–71. https://doi.org/10.1128/aac.01410-08.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Livermore DM. β-Lactamases in laboratory and clinical resistance. Clin Microbiol Rev. 1995;8(4):557–84. https://doi.org/10.1128/cmr.8.4.557.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Meini S, Tascini C, Cei M, Sozio E, Rossolini GM. AmpC β-lactamase-producing Enterobacterales: what a clinician should know. Infection. 2019;47(3):363–75. https://doi.org/10.1007/s15010-019-01291-9.

    Article  CAS  PubMed  Google Scholar 

  6. Tamma PD, Doi Y, Bonomo RA, Johnson JK, Simner PJ. A Primer on AmpC β-Lactamases: Necessary Knowledge for an Increasingly Multidrug-resistant World. Clin Infect Dis. 2019;69(8):1446–55. https://doi.org/10.1093/cid/ciz173.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Carlier M, Carrette S, Roberts JA, Stove V, Verstraete A, Hoste E, et al. Meropenem and piperacillin/tazobactam prescribing in critically ill patients: does augmented renal clearance affect pharmacokinetic/pharmacodynamic target attainment when extended infusions are used? Crit Care. 2013;17(3):R84. https://doi.org/10.1186/cc12705.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Udy AA, Varghese JM, Altukroni M, Briscoe S, McWhinney BC, Ungerer JP, et al. Subtherapeutic initial β-lactam concentrations in select critically ill patients: association between augmented renal clearance and low trough drug concentrations. Chest. 2012;142(1):30–9. https://doi.org/10.1378/chest.11-1671.

    Article  CAS  PubMed  Google Scholar 

  9. Centers for Disease Control and Prevention. Antibiotic resistance threats in the United States. 2019. https://www.cdc.gov/drugresistance/pdf/threats-report/2019-ar-threats-report-508.pdf. Accessed 23 Feb 2022 .

    Book  Google Scholar 

  10. Motsch J, Murta de Oliveira C, Stus V, Köksal I, Lyulko O, Boucher HW, et al. RESTORE-IMI 1: a multicenter, randomized, double-blind trial comparing efficacy and safety of imipenem/relebactam vs colistin plus imipenem in patients with imipenem-nonsusceptible bacterial infections. Clin Infect Dis. 2020;70(9):1799–808. https://doi.org/10.1093/cid/ciz530.

    Article  CAS  PubMed  Google Scholar 

  11. World Health Organization. Global action plan on antimicrobial resistance. 2015. https://www.amcra.be/swfiles/files/WHO%20actieplan_90.pdf. Accessed 23 March 2022.

    Google Scholar 

  12. Titov I, Wunderink RG, Roquilly A, Rodríguez Gonzalez D, David-Wang A, Boucher HW, et al. A randomized, double-blind, multicenter trial comparing efficacy and safety of imipenem/cilastatin/relebactam versus piperacillin/tazobactam in adults with hospital-acquired or ventilator-associated bacterial pneumonia (RESTORE-IMI 2 study). Clin Infect Dis. 2021;73(11):e4539–48. https://doi.org/10.1093/cid/ciaa803.

    Article  PubMed  Google Scholar 

  13. Roberts JA, Nicolau DP, Martin-Loeches I, DeRyke CA, Losada MC, Du J, et al. Imipenem/cilastatin/relebactam efficacy, safety, and probability of target attainment in adults with hospital-acquired or ventilator-associated bacterial pneumonia among patients with baseline renal impairment, normal renal function, and augmented renal clearance. 2022.

  14. RECARBRIO™ (imipenem, cilastatin, and relebactam), for injection, for intravenous use. Prescribing information. Rahway: Merck Sharp & Dohme LLC; 2022.

  15. MSD. Recarbrio: EU summary of product characteristics. 2021. https://www.ema.europa.eu. Accessed 21 April 2022.

    Google Scholar 

  16. Young K, Painter RE, Raghoobar SL, Hairston NN, Racine F, Wisniewski D, et al. In vitro studies evaluating the activity of imipenem in combination with relebactam against Pseudomonas aeruginosa. BMC Microbiol. 2019;19(1):150. https://doi.org/10.1186/s12866-019-1522-7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Morrissey I, Hackel M, Badal R, Bouchillon S, Hawser S, Biedenbach D. A review of ten years of the Study for Monitoring Antimicrobial Resistance Trends (SMART) from 2002 to 2011. Pharmaceuticals (Basel). 2013;6(11):1335–46. https://doi.org/10.3390/ph6111335.

    Article  CAS  PubMed  Google Scholar 

  18. Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing. 31st ed. CLSI supplement M100. 31st ed. Wayne: Clinical Laboratory Standards Institute; 2021.

    Google Scholar 

  19. Quale J, Bratu S, Gupta J, Landman D. Interplay of efflux system, ampC, and oprD expression in carbapenem resistance of Pseudomonas aeruginosa clinical isolates. Antimicrob Agents Chemother. 2006;50(5):1633–41. https://doi.org/10.1128/aac.50.5.1633-1641.2006.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Bonomo RA, Burd EM, Conly J, Limbago BM, Poirel L, Segre JA, et al. Carbapenemase-Producing Organisms: A Global Scourge. Clin Infect Dis. 2018;66(8):1290–7. https://doi.org/10.1093/cid/cix893.

    Article  CAS  PubMed  Google Scholar 

  21. Livermore DM. Interplay of impermeability and chromosomal β-lactamase activity in imipenem-resistant Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1992;36(9):2046–8. https://doi.org/10.1128/aac.36.9.2046.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Horner C, Mushtaq S, Livermore DM. Potentiation of imipenem by relebactam for Pseudomonas aeruginosa from bacteraemia and respiratory infections. J Antimicrob Chemother. 2019;74(7):1940–4. https://doi.org/10.1093/jac/dkz133.

    Article  CAS  PubMed  Google Scholar 

  23. EUCAST. Redefining susceptibility testing categories S, I and R. 2019. https://www.eucast.org/newsiandr. Accessed 30 Sept 2022 .

    Google Scholar 

  24. Tamma PD, Aitken SL, Bonomo RA, Mathers AJ, van Duin D, Clancy CJ. Infectious Diseases Society of America 2022 guidance on the treatment of extended-spectrum β-lactamase producing Enterobacterales (ESBL-E), carbapenem-resistant Enterobacterales (CRE), and Pseudomonas aeruginosa with difficult-to-treat resistance (DTR-P. aeruginosa). Clin Infect Dis. 2022;75(2):187–212. https://doi.org/10.1093/cid/ciac268.

    Article  PubMed  PubMed Central  Google Scholar 

  25. Guilhaumou R, Benaboud S, Bennis Y, Dahyot-Fizelier C, Dailly E, Gandia P, et al. Optimization of the treatment with beta-lactam antibiotics in critically ill patients-guidelines from the French Society of Pharmacology and Therapeutics (Société Française de Pharmacologie et Thérapeutique-SFPT) and the French Society of Anaesthesia and Intensive Care Medicine (Société Française d’Anesthésie et Réanimation-SFAR). Crit Care. 2019;23(1):104. https://doi.org/10.1186/s13054-019-2378-9.

    Article  PubMed  PubMed Central  Google Scholar 

  26. Lala M, Brown M, Kantesaria B, Walker B, Paschke A, Rizk ML. Simplification of imipenem dosing by removal of weight-based adjustments. J Clin Pharmacol. 2019;59(5):646–53. https://doi.org/10.1002/jcph.1356.

    Article  CAS  PubMed  Google Scholar 

  27. Ambrose PG, Bhavnani SM, Rubino CM, Louie A, Gumbo T, Forrest A, et al. Pharmacokinetics-pharmacodynamics of antimicrobial therapy: it’s not just for mice anymore. Clin Infect Dis. 2007;44(1):79–86. https://doi.org/10.1086/510079.

    Article  CAS  PubMed  Google Scholar 

  28. Patel M, Daryani NM, Feng H-P, Hilbert DW, Melchers MJ, Mavridou E, et al.: Imipenem/relebactam pharmacokinetic/pharmacodynamic analyses from an in vivo neutropenic murine thigh infection model. In: European Congress of Clinical Microbiology and Infectious Diseases (ECCMID). Paris, France; 2020: Abstract 1693.

  29. Daryani N, Patel M, Flattery A, Young K, Rizk ML: Imipenem/relebactam pharmacokinetic/pharmacodynamic analyses from an in vivo neutropenic mouse delayed lung infection model. In: European Congress of Clinical Microbiology and Infectious Diseases (ECCMID). Paris, France; 2020: Abstract 2086.

  30. Bhagunde P, Zhang Z, Racine F, Carr D, Wu J, Young K, et al. A translational pharmacokinetic/pharmacodynamic model to characterize bacterial kill in the presence of imipenem-relebactam. Int J Infect Dis. 2019;89:55–61. https://doi.org/10.1016/j.ijid.2019.08.026.

    Article  CAS  PubMed  Google Scholar 

  31. Wu J, Racine F, Wismer MK, Young K, Carr DM, Xiao JC, et al. Exploring the pharmacokinetic/pharmacodynamic relationship of relebactam (MK-7655) in combination with imipenem in a hollow-fiber infection model. Antimicrob Agents Chemother. 2018;62(5):e02323-e2417. https://doi.org/10.1128/aac.02323-17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Bhagunde P, Patel P, Lala M, Watson K, Copalu W, Xu M, et al. Population pharmacokinetic analysis for imipenem-relebactam in healthy volunteers and patients with bacterial infections. CPT Pharmacometrics Syst Pharmacol. 2019;8(10):748–58. https://doi.org/10.1002/psp4.12462.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Patel M, Bellanti F, Daryani NM, Noormohamed N, Hilbert DW, Young K, et al. Population pharmacokinetic/pharmacodynamic assessment of imipenem/cilastatin/relebactam in patients with hospital-acquired/ventilator-associated bacterial pneumonia. Clin Transl Sci. 2022;15(2):396–408. https://doi.org/10.1111/cts.13158.

    Article  CAS  PubMed  Google Scholar 

  34. Hirvonen VHA, Spencer J, van der Kamp MW. Antimicrobial Resistance Conferred by OXA-48 β-Lactamases: Towards a Detailed Mechanistic Understanding. Antimicrob Agents Chemother. 2021;65(6); https://doi.org/10.1128/aac.00184-21.

  35. Boyd SE, Holmes A, Peck R, Livermore DM, Hope W. OXA-48-Like β-Lactamases: Global Epidemiology, Treatment Options, and Development Pipeline. Antimicrob Agents Chemother. 2022;66(8):e0021622. https://doi.org/10.1128/aac.00216-22.

    Article  CAS  PubMed  Google Scholar 

  36. Wen X, Gehring R, Stallbaumer A, Riviere JE, Volkova VV. Limitations of MIC as sole metric of pharmacodynamic response across the range of antimicrobial susceptibilities within a single bacterial species. Sci Rep. 2016;6:37907. https://doi.org/10.1038/srep37907.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Piddock LJ, Wise R. Newer mechanisms of resistance to beta-lactam antibiotics in gram-negative bacteria. J Antimicrob Chemother. 1985;16(3):279–84. https://doi.org/10.1093/jac/16.3.279.

    Article  CAS  PubMed  Google Scholar 

  38. Yao Y, Falgenhauer L, Falgenhauer J, Hauri AM, Heinmüller P, Domann E, et al. Carbapenem-resistant Citrobacter spp. as an emerging concern in the hospital-setting: results from a genome-based regional surveillance study. Front Cell Infect Microbiol. 2021;11:744431. https://doi.org/10.3389/fcimb.2021.744431.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Clinical and Laboratory Standards Institute. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically. 11th ed. CLSI supplement M07. Wayne: Clinical Laboratory Standards Institute; 2018.

    Google Scholar 

  40. Lob SH, Kazmierczak KM, Badal RE, Hackel MA, Bouchillon SK, Biedenbach DJ, et al. Trends in susceptibility of Escherichia coli from intra-abdominal infections to ertapenem and comparators in the United States according to data from the SMART program, 2009 to 2013. Antimicrob Agents Chemother. 2015;59(6):3606–10. https://doi.org/10.1128/AAC.05186-14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Nichols WW, de Jonge BL, Kazmierczak KM, Karlowsky JA, Sahm DF. In vitro susceptibility of global surveillance isolates of Pseudomonas aeruginosa to ceftazidime-avibactam (INFORM 2012 to 2014). Antimicrob Agents Chemother. 2016;60(8):4743–9. https://doi.org/10.1128/AAC.00220-16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Estabrook M, Kazmierczak KM, Wise M, Arhin FF, Stone GG, Sahm DF. Molecular characterization of clinical isolates of Enterobacterales with elevated MIC values for aztreonam-avibactam from the INFORM global surveillance study, 2012–2017. J Glob Antimicrob Resist. 2021;24:316–20. https://doi.org/10.1016/j.jgar.2021.01.010.

    Article  CAS  PubMed  Google Scholar 

  43. Bortolaia V, Kaas RS, Ruppe E, Roberts MC, Schwarz S, Cattoir V, et al. ResFinder 4.0 for predictions of phenotypes from genotypes. J Antimicrob Chemother. 2020;75(12):3491–500. https://doi.org/10.1093/jac/dkaa345.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

Medical writing and/or editorial assistance was provided Rebecca L. Orndorff, PhD, CMPP, and Meredith Rogers, MS, CMPP, of The Lockwood Group, Stamford, CT, USA. This assistance was funded by Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA (MSD). This research was presented virtually in part at the 31st European Congress of Clinical Microbiology and Infectious Diseases, July 9–12, 2021.

Funding

Funding for this research was provided by Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

Author information

Authors and Affiliations

Authors

Contributions

All authors are responsible for the work described in this paper and meet ICMJE authorship criteria. All authors were involved in at least one of the following: conception (DH, CAD, KY), design of work or acquisition (CAD, MH, KY), analysis (DH, CAD, KY), interpretation of data (DH, CAD, MM, KY), and drafting the manuscript and/or revising/reviewing the manuscript for important intellectual content (DH, CAD, MM, MH, KY). All authors provided final approval of the version to be published. All authors agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Corresponding author

Correspondence to David W. Hilbert.

Ethics declarations

Ethics approval and consent to participate

All methods were carried out in accordance with the ethical principles of the Declaration of Helsinki and all relevant guidelines and regulations. Ethical approval and informed consent were not required because all isolates received into the study followed multiple subcultures and were completely de-identified. The secondary research use of de-identified isolates is considered exempt research according to the Regulations for the Protection of Human Subjects in Research of the U.S. Department of Health and Human Services, Office for Human Research Protections (45 CFR 46).

Consent for publication

Not applicable.

Competing interests

DWH, CAD, MM, and KY are employees of Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA (MSD), who may own stock and/or hold stock options in Merck & Co., Inc., Rahway, NJ, USA. MH is an employee of IHMA, Inc., Schaumburg, IL, USA. IHMA received financial support from MSD to perform the analyses presented.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Additional file 1.

Molecular characterization of resistance mechanisms on the subset of samples with data available.

Additional file 2.

Relebactam enhances the activity of imipenem among Enterobacterales.

Additional file 3.

Imipenem/relebactam susceptibility of imipenem nonsusceptible isolates by region.

Additional file 4.

  Graphical abstract.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Hilbert, D.W., DeRyke, C.A., Motyl, M. et al. Relebactam restores susceptibility of resistant Pseudomonas aeruginosa and Enterobacterales and enhances imipenem activity against chromosomal AmpC-producing species: analysis of global SMART 2018–2020. BMC Microbiol 23, 165 (2023). https://doi.org/10.1186/s12866-023-02864-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s12866-023-02864-3

Keywords