Broad-spectrum Β-lactamase Research Articles

In-vitro activity of ceftolozane/tazobactam against Pseudomonas aeruginosa and Enterobacterales isolates collected from two general hospitals in Singapore.

The emerging resistance to cephalosporins and carbapenems in gram-negative pathogens poses significant health challenges and increased treatment failures. The development and evaluation of novel therapeutic options are needed urgently. This study aimed to assess the in vitro activity of ceftolozane/tazobactam (C/T), a new cephalosporin and β-lactamase inhibitor combination, against isolates of Pseudomonas aeruginosa, Escherichia coli, and Klebsiella pneumoniae from different infection sources in two general hospitals in Singapore. Susceptibility testing revealed that C/T has good activity against 600 tested gram-negative pathogens, showing a 94.8% susceptibility rate across different species and infection types. Additionally, emergence of C/T resistance was also observed and characterized with whole genome sequencing (WGS) analysis. WGS analysis indicated that C/T resistance was predominantly associated with harbouring broad-spectrum β-lactamases such as carbapenemases, ESBLs, AmpC and others. Novel mutations in penicillin-binding-protein 3 (PBP3), the target of ceftolozane, and variants associated with hyper ampC expression (ampD, ampR, and dacB), were also found in C/T-resistant isolates, potentially contributing to C/T resistance. The finding of this study provides valuable local data on C/T susceptibility, which is important for guiding the proper clinical use of C/T and promoting antimicrobial stewardship in managing gram-negative infections.

P-1056. Association of High-Risk Clones Of P. aeruginosa With MBLs Poses Heightened Treatment Challenge, however there is a Silver Lining

Abstract Background Globally, ‘high-risk’ clones of P. aeruginosa continue to pose treatment challenges. These clones, in addition to intrinsic non-enzymatic β-lactam resistance mechanisms such as hyper-expressing efflux and down-regulated porins, have been acquiring broad-spectrum β-lactamases including carbapenemases. In India, recent reports indicate that most of carbapenem-resistant isolates carry MBLs in particular NDMs. However, genomic characterization of such isolates is lacking and therefore their relatedness to internationally circulating high-risk clones is not known. Herein, we have genomically characterized 35 P. aeruginosa index isolates collected from hospitalised patients including admitted at our intensive care unit in 2022 - 2024. We also determined their susceptibility to cefepime/zidebactam (Phase 3-stage) and other newer BL/BLIs, for both serine and MBL-producing Gram-negative bacteria. Methods The index P. aeruginosa pathogens were isolated from sputum, tracheal secretion, blood and drain fluid and identified by Vitek 2 (one isolate per patient). The whole genome of these isolates were sequenced (WGS) using HiSeq (Illumina 6000 platform). Susceptibilities to cefepime/zidebactam and other antibiotics were determined by reference broth micro-dilution MIC method according to CLSI M07-A11, 2018. Results WGS revealed existence of 9 distinct multi-locus sequence types (ST), a high clonal variation despite isolates belonged to a single centre (Figure 1). Among them, 3 were international high-risk clones and nearly half of the isolated belonged to these 3 STs. Three MBL types, NDM, VIM and DIM were detected and none of the isolates carried serine carbapenemase. Mutations were noted in genes encoding efflux and impermeability. Susceptibilities to aminoglycosides and fluoroquinolones were < 30%. Except 5, all isolates were resistant to carbapenems. There was a 100% susceptibility to cefepime/zidebactam Conclusion WGS analysis showed multiplicity of resistance mechanisms including MBLs in high-risk clones thus severely limiting treatment options. Investigational antibiotic cefepime/zidebactam may be a potential treatment option for infections caused by such isolates. In India VIM was the most common Carbapenemase resistance is being replaced by MBL(NDM) Disclosures All Authors: No reported disclosures

Relative inhibitory activities of newly developed diazabicyclooctanes, boronic acid derivatives, and penicillin-based sulfone β-lactamase inhibitors against broad-spectrum AmpC β-lactamases.

The relative inhibitory activities of diazabicyclooctanes (avibactam, relebactam, zidebactam, nacubactam, durlobactam), boronic acid derivatives (vaborbactam, taniborbactam, xeruborbactam), and penicillin-based sulfone derivative enmetazobactam were evaluated against several intrinsic and acquired class C β-lactamases. By contrast to vaborbactam and enmetazobactam, taniborbactam, xeruborbactam, and all diazabicyclooctanes demonstrated effective activities against most AmpC enzymes. Notably, durlobactam exhibited the most pronounced inhibitory effect. Interstingly, the chromosomal AmpC of Acinetobacter baumannii was the least sensitive enzyme to the newly developed β-lactamase inhibitors.

Cefepime-taniborbactam demonstrates potent in vitro activity vs Enterobacterales with blaOXA-48.

Taniborbactam (formerly VNRX-5133) is a novel, investigational boronic acid β-lactamase inhibitor. The combination of cefepime (FEP) with taniborbactam is active against Enterobacterales carrying class A, B, C, and/or D enzymes. We assessed the activity of FEP-taniborbactam against Enterobacterales clinical strains carrying blaOXA-48 (N = 50, 100%), of which 78% harbored at least one extended-spectrum β-lactamase (ESBL). CLSI-based agar dilution susceptibility testing was conducted using FEP-taniborbactam and comparators FEP, meropenem-vaborbactam (MVB), and ceftazidime-avibactam (CZA). The addition of taniborbactam lowered FEP MICs to the provisionally susceptible range of ≤16 µg/mL; the MIC90 value decreased from ≥64 µg/mL for FEP to 4 µg/mL for FEP-taniborbactam. Notably, FEP-taniborbactam MIC50/MIC90 values (0.5/4 µg/mL) were lower than those for MVB (1/16 µg/mL) and comparable to those for CZA (0.5/1 µg/mL). Time-kill assays with E. coli clinical strains DOV (blaOXA-48, blaCTX-M-15, blaTEM-1, and blaOXA-1) and MLI (blaOXA-48, blaVEB, blaTEM-1, and blaCMY-2) revealed that FEP-taniborbactam at concentrations 1×, 2×, and 4× MIC displayed time-dependent reductions in the number of CFU/mL from 0 to 6 h, and at 4× MIC demonstrated bactericidal activity (3 log10 reduction in CFU/mL at 24 h). Therefore, taniborbactam in combination with FEP was highly active against this diverse panel of Enterobacterales with blaOXA-48 and represents a potential addition to our antibiotic arsenal.IMPORTANCEOXA-48-like β-lactamases are class D carbapenemases widespread in Klebsiella pneumoniae and other Enterobacterales and are associated with carbapenem treatment failures. As up to 80% of OXA-48-like positive isolates coproduce extended-spectrum β-lactamases, a combination of β-lactams with broad-spectrum β-lactamase inhibitors is required to counteract all OXA-48-producing strains effectively. Herein, we evaluated the activity of cefepime-taniborbactam against 50 clinical strains producing OXA-48. We report that adding taniborbactam shifted the minimum inhibitory concentration (MIC) toward cefepime's susceptible range, restoring its antimicrobial activity. Notably, cefepime-taniborbactam MIC50/MIC90 values (0.5/4 µg/mL) were comparable to ceftazidime-avibactam (0.5/1 µg/mL). Finally, time-kill assays revealed sustained bactericidal activity of cefepime-taniborbactam for up to 24 h. In conclusion, cefepime-taniborbactam will be a welcome addition to the antibiotic arsenal to combat Enterobacterales producing OXA-48.

Convergence of mcr-1 and broad-spectrum β-lactamase genes in Escherichia coli strains from the environmental sector

The mcr-type gene encodes the main plasmid-mediated mechanism of colistin resistance and has been reported in several bacterial species obtained from different sources. Anthropogenic activities in the environment favor the evolution of antimicrobial resistance. Indeed, mcr-1-positive Escherichia coli strains were susceptible to non-polymyxins antimicrobials, but now emerging as multidrug-resistant (MDR) lineages. In this regard, hundreds of surface water and agricultural soil samples were screened for the presence of E. coli carrying the mcr-type genes and mcr-1-positive strains were subjected to in-depth genomic analysis. Almost all colistin-resistant strains were classified as MDR, highlighting those obtained from soils that showed resistance to extended-spectrum cephalosporins and carbapenems. International and high-risk clones of E. coli were identified, with ST10 and ST1720 shared between water and soil samples. Resistome analysis showed a broad resistome (AMR, metal tolerance, and biocide resistance). The mcr-1.1 and mcr-1.26 allelic variants were detected on IncX4 and IncI2 plasmids. Curiously, mcr-1-positive E. coli strains from agricultural soils harbored plasmid-mediated blaCTX-M-1, blaCTX-M-8, or blaKPC-2 genes. Virulome analysis demonstrated traits of a high putative virulence potential, with the presence of extraintestinal pathogenic E. coli. Comparative analysis revealed the persistence and dissemination of plasmid-mediated antimicrobial resistance genes in genetically diversity E. coli strains at the human-animal-environmental interface. These findings demonstrate a possible emerging AMR trend with the convergence of resistance to colistin and broad-spectrum β-lactams in environmental-derived E. coli strains.

In vitro and intracellular activity of vaborbactam combined with β-lactams against Mycobacterium abscessus.

Mycobacterium abscessus has emerged as an opportunistic pathogen responsible for lung infections, especially in cystic fibrosis patients. In spite of the production of the broad-spectrum β-lactamase BlaMab, the carbapenem imipenem is recommended in the initial phase of the treatment of pulmonary infections. Here, we determine whether the addition of vaborbactam, a second-generation β-lactamase inhibitor belonging to the boronate family, improves the activity of β-lactams against M. abscessus. The activity of β-lactams, alone or in combination with vaborbactam, was evaluated against M. abscessus CIP104536 by determining MICs, time-killing and intramacrophage activity. Kinetic parameters for the inhibition of BlaMab by vaborbactam were determined by spectrophotometry. The combination of vaborbactam (8 mg/L) with β-lactams decreased more than 8 times the MIC of amoxicillin (from >1024 to 128 mg/L) and 2 times the MICs of meropenem (from 16 to 8 mg/L) and imipenem (from 4 to 2 mg/L). The reduction of the MICs was less than that obtained with avibactam at 4 mg/L for amoxicillin (from >1024 to 16 mg/L, more than 64 times less) and for meropenem (from 16 to 4 mg/L, 4 times less). In vitro and intracellularly, M. abscessus was not killed by the meropenem/vaborbactam combination, in spite of significant in vitro inhibition of BlaMab by vaborbactam. Inhibition of BlaMab by vaborbactam decreases the MIC of β-lactams, including that of meropenem. As meropenem/vaborbactam is clinically available, this combination offers an alternative therapeutic option that should be evaluated for the treatment of pulmonary infections due to M. abscessus.

In vitro and in vivo activities of KSP-1007, a broad-spectrum inhibitor of serine- and metallo-β-lactamases, in combination with meropenem against carbapenem-resistant Gram-negative bacteria.

KSP-1007 is a novel bicyclic boronate-based broad-spectrum β-lactamase inhibitor and is being developed in combination with meropenem (MEM) for the treatment of infections caused by carbapenem-resistant Gram-negative bacteria, a global health concern, and here, we describe its characteristics. KSP-1007 exhibited low apparent inhibition constant (Ki app) values against all classes of β-lactamase, including imipenemase types and oxacillinase types from Acinetobacter baumannii. Against 207 Enterobacterales and 55 A. baumannii, including carbapenemase producers, KSP-1007 at fixed concentrations of 4, 8, and 16 µg/mL dose-dependently potentiated the in vitro activity of MEM in broth microdilution MIC testing. The MIC90 of MEM/KSP-1007 at 8 µg/mL against Enterobacterales was lower than those of MEM/vaborbactam, ceftazidime/avibactam, imipenem/relebactam, and colistin and similar to those of aztreonam/avibactam, cefiderocol, and tigecycline. The in vitro activity of MEM/KSP-1007 at ≥4 µg/mL against Enterobacterales harboring metallo-β-lactamase was superior to that of cefepime/taniborbactam. MEM/KSP-1007 showed excellent activity against Escherichia coli with PBP3 mutations and New Delhi metallo-β-lactamase compared to aztreonam/avibactam, cefepime/taniborbactam, and cefiderocol. MEM/KSP-1007 at 8 µg/mL showed greater efficacy against A. baumannii than these comparators except for cefiderocol, tigecycline, and colistin. A 2-fold reduction in MEM MIC against 96 Pseudomonas aeruginosa was observed in combination with KSP-1007. MEM/KSP-1007 demonstrated bactericidal activity against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa based on minimum bactericidal concentration/MIC ratios of ≤4. KSP-1007 enhanced the in vivo activity of MEM against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa in murine systemic, complicated urinary tract, and thigh infection models. Collectively, MEM/KSP-1007 has a good profile for treating carbapenem-resistant Gram-negative bacterial infections.

Relative inhibitory activities of the broad-spectrum β-lactamase inhibitor xeruborbactam in comparison with taniborbactam against metallo-β-lactamases produced in Escherichia coli and Pseudomonas aeruginosa.

Xeruborbactam is a newly developed β-lactamase inhibitor designed for metallo-β-lactamases (MBLs). This study assessed the relative inhibitory properties of this novel inhibitor in comparison with another MBL inhibitor, namely taniborbactam (TAN), against a wide range of acquired MBL produced either in Escherichia coli or Pseudomonas aeruginosa. As observed with taniborbactam, the combination of xeruborbactam (XER) with β-lactams, namely, ceftazidime, cefepime and meropenem, led to significantly decreased MIC values for a wide range of B1-type MBL-producing E. coli, including most recombinant strains producing NDM, VIM, IMP, GIM-1, and DIM-1 enzymes. Noteworthily, while TAN-based combinations significantly reduced MIC values of β-lactams for MBL-producing P. aeruginosa recombinant strains, those with XER were much less effective. We showed that this latter feature was related to the MexAB-OprM efflux pump significantly impacting MIC values when testing XER-based combinations in P. aeruginosa. The relative inhibitory concentrations (IC50 values) were similar for XER and TAN against NDM and VIM enzymes. Noteworthily, XER was effective against NDM-9, NDM-30, VIM-83, and most of IMP enzymes, although those latter enzymes were considered resistant to TAN. However, no significant inhibition was observed with XER against IMP-10, SPM-1, and SIM-1 as well as the representative subclass B2 and B3 enzymes, PFM-1 and AIM-1. The determination of the constant inhibition (Ki) of XER revealed a much higher value against IMP-10 than against NDM-1, VIM-2, and IMP-1. Hence, IMP-10 that differs from IMP-1 by a single amino-acid substitution (Val67Phe) can, therefore, be considered resistant to XER.

3-O-Substituted Quercetin: an Antibiotic-Potentiating Agent against Multidrug-Resistant Gram-Negative Enterobacteriaceae through Simultaneous Inhibition of Efflux Pump and Broad-Spectrum Carbapenemases.

The discovery of safe and efficient inhibitors against efflux pumps as well as metallo-β-lactamases (MBL) is one of the main challenges in the development of multidrug-resistant (MDR) reversal agents which can be utilized in the treatment of carbapenem-resistant Gram-negative bacteria. In this study, we have identified that introduction of an ethylene-linked sterically demanding group at the 3-OH position of the previously reported MDR reversal agent di-F-Q endows the resulting compounds with hereto unknown multitarget inhibitory activity against both efflux pumps and broad-spectrum β-lactamases including difficult-to-inhibit MBLs. A molecular docking study of the multitarget inhibitors against efflux pump, as well as various classes of β-lactamases, revealed that the 3-O-alkyl substituents occupy the novel binding sites in efflux pumps as well as carbapenemases. Not surprisingly, the multitarget inhibitors rescued the antibiotic activity of a carbapenem antibiotic, meropenem (MEM), in NDM-1 (New Delhi Metallo-β-lactamase-1)-producing carbapenem-resistant Enterobacteriaceae (CRE), and they reduced MICs of MEM more than four-fold (synergistic effect) in 8-9 out of 14 clinical strains. The antibiotic-potentiating activity of the multitarget inhibitors was also demonstrated in CRE-infected mouse model. Taken together, these results suggest that combining inhibitory activity against two critical targets in MDR Gram-negative bacteria, efflux pumps, and β-lactamases, in one molecule is possible, and the multitarget inhibitors may provide new avenues for the discovery of safe and efficient MDR reversal agents.

When does antimicrobial resistance increase bacterial fitness? Effects of dosing, social interactions, and frequency dependence on the benefits of AmpC β-lactamases in broth, biofilms, and a gut infection model.

One of the longstanding puzzles of antimicrobial resistance is why the frequency of resistance persists at intermediate levels. Theoretical explanations for the lack of fixation of resistance include cryptic costs of resistance or negative frequency-dependence but are seldom explored experimentally. β-lactamases, which detoxify penicillin-related antibiotics, have well-characterized frequency-dependent dynamics driven by cheating and cooperation. However, bacterial physiology determines whether β-lactamases are cooperative, and we know little about the sociality or fitness of β-lactamase producers in infections. Moreover, media-based experiments constrain how we measure fitness and ignore important parameters such as infectivity and transmission among hosts. Here, we investigated the fitness effects of broad-spectrum AmpC β-lactamases in Enterobacter cloacae in broth, biofilms, and gut infections in a model insect. We quantified frequency- and dose-dependent fitness using cefotaxime, a third-generation cephalosporin. We predicted that infection dynamics would be similar to those observed in biofilms, with social protection extending over a wide dose range. We found evidence for the sociality of β-lactamases in all contexts with negative frequency-dependent selection, ensuring the persistence of wild-type bacteria, although cooperation was less prevalent in biofilms, contrary to predictions. While competitive fitness in gut infections and broth had similar dynamics, incorporating infectivity into measurements of fitness in infections significantly affected conclusions. Resistant bacteria had reduced infectivity, which limited the fitness benefits of resistance to infections challenged with low antibiotic doses and low initial frequencies of resistance. The fitness of resistant bacteria in more physiologically tolerant states (in biofilms, in infections) could be constrained by the presence of wild-type bacteria, high antibiotic doses, and limited availability of β-lactamases. One conclusion is that increased tolerance of β-lactams does not necessarily increase selection pressure for resistance. Overall, both cryptic fitness costs and frequency dependence curtailed the fitness benefits of resistance in this study.

Durlobactam, a Diazabicyclooctane β-Lactamase Inhibitor, Inhibits BlaC and Peptidoglycan Transpeptidases of Mycobacterium tuberculosis.

Peptidoglycan synthesis is an underutilized drug target in Mycobacterium tuberculosis (Mtb). Diazabicyclooctanes (DBOs) are a class of broad-spectrum β-lactamase inhibitors that also inhibit certain peptidoglycan transpeptidases that are important in mycobacterial cell wall synthesis. We evaluated the DBO durlobactam as an inhibitor of BlaC, the Mtb β-lactamase, and multiple Mtb peptidoglycan transpeptidases (PonA1, LdtMt1, LdtMt2, LdtMt3, and LdtMt5). Timed electrospray ionization mass spectrometry (ESI-MS) captured acyl-enzyme complexes with BlaC and all transpeptidases except LdtMt5. Inhibition kinetics demonstrated durlobactam was a potent and efficient DBO inhibitor of BlaC (KIapp 9.2 ± 0.9 μM, k2/K 5600 ± 560 M-1 s-1) and similar to clavulanate (KIapp 3.3 ± 0.6 μM, k2/K 8400 ± 840 M-1 s-1); however, durlobactam had a lower turnover number (tn = kcat/kinact) than clavulanate (1 and 8, respectively). KIapp values with durlobactam and clavulanate were similar for peptidoglycan transpeptidases, but ESI-MS captured durlobactam complexes at more time points. Molecular docking and simulation demonstrated several productive interactions of durlobactam in the active sites of BlaC, PonA1, and LdtMt2. Antibiotic susceptibility testing was conducted on 11 Mtb isolates with amoxicillin, ceftriaxone, meropenem, imipenem, clavulanate, and durlobactam. Durlobactam had a minimum inhibitory concentration (MIC) range of 0.5-16 μg/mL, similar to the ranges for meropenem (1-32 μg/mL) and imipenem (0.5-64 μg/mL). In β-lactam + durlobactam combinations (1:1 mass/volume), MICs were lowered 4- to 64-fold for all isolates except one with meropenem-durlobactam. This work supports further exploration of novel β-lactamase inhibitors that target BlaC and Mtb peptidoglycan transpeptidases.

Herbal Products-Powered Thermosensitive Hydrogel with Phototherapy and Microenvironment Reconstruction for Accelerating Multidrug-Resistant Bacteria-Infected Wound Healing.

Wound healing and infection remain significant challenges due to the ineffectiveness against multidrug-resistant (MDR) bacteria and the complex oxidative wound microenvironments. To address these issues, thymoquinone-reinforced injectable and thermosensitive TQ@PEG-PAF-Cur hydrogels with dual functions of microenvironment reshaping and photodynamic therapy are developed. The hydrogel comprises natural compound thymoquinone (TQ) and poly (ethylene glycol)-block-poly (alanine-co-phenyl alanine) copolymers (PEG-PAF) conjugated with natural photosensitizer curcumin (Cur). The incorporation of TQ and Cur reduces the sol-to-gel transition temperature of TQ@PEG-PAF-Cur to 30°C, compared to PEG-PAF hydrogel (37°C), due to the formation of strong hydrogen bonding, matching the wound microenvironment temperature. Under blue light excitation, TQ@PEG-PAF-Cur generates significant amounts of reactive oxygen species such as H2O2, 1O2, and ·OH, exhibiting rapid and efficient bactericidal capacities against methicillin-resistant Staphylococcus aureus and broad spectrum β-lactamases Escherichia coli via photodynamic therapy (PDT). Additionally, Cur effectively inhibits the expressions of proinflammatory cytokines in skin tissue-forming cells. As a result, the composite hydrogel can rapidly transform into a gel to cover the wound, reshape the wound microenvironment, and accelerate wound healing in vivo. This collaborative antibacterial strategy provides valuable insights to guide the development of multifunctional materials for efficient wound healing.

Coexistence of blaIMP−4 and blaSFO−1 in an IncHI5B plasmid harbored by tigecycline-non-susceptible Klebsiella variicola strain

BackgroundKlebsiella variicola is considered a newly emerging human pathogen. Clinical isolates of carbapenemase and broad-spectrum β-lactamase-producing K. variicola remain relatively uncommon. A strain of K. variicola 4253 was isolated from a clinical sample, and was identified to carry the blaIMP−4 and blaSFO−1 genes. This study aims to discern its antibiotic resistance phenotype and genomic characteristics.MethodsSpecies identification was conducted using MALDI-TOF/MS. PCR identification confirmed the presence of the blaIMP−4 and blaSFO−1 genes. Antibiotic resistance phenotype and genomic characteristics were detected by antimicrobial susceptibility testing and whole-genome sequencing. Plasmid characterization was carried out through S1-PFGE, conjugation experiments, Southern blot, and comparative genomic analysis.ResultsK. variicola 4253 belonged to ST347, and demonstrated resistance to broad-spectrum β-lactamase drugs and tigecycline while being insensitive to imipenem and meropenem. The blaIMP−4 and blaSFO−1 genes harbored on the plasmid p4253-imp. The replicon type of p4253-imp was identified as IncHI5B, representing a multidrug-resistant plasmid capable of horizontal transfer and mediating the dissemination of drug resistance. The blaIMP−4 gene was located on the In809-like integrative element (Intl1-blaIMP−4-aacA4-catB3), which circulates in Acinetobacter and Enterobacteriaceae.ConclusionsThis study reports the presence of a strain of K. variicola, which is insensitive to tigecycline, carrying a plasmid harboring blaIMP−4 and blaSFO−1. It is highly likely that the strain acquired this plasmid through horizontal transfer. The blaIMP−4 array (Intl1-blaIMP−4-aacA4-catB3) is also mobile in Acinetobacter and Enterobacteriaceae. So it is essential to enhance clinical awareness and conduct epidemiological surveillance on multidrug-resistant K. variicola, conjugative plasmids carrying blaIMP−4, and the In809 integrative element.

Effect of modification of penicillin-binding protein 3 on susceptibility to ceftazidime-avibactam, imipenem-relebactam, meropenem-vaborbactam, aztreonam-avibactam, cefepime-taniborbactam, and cefiderocol of Escherichia coli strains producing broad-spectrum β-lactamases.

The impact of penicillin-binding protein 3 (PBP3) modifications that may be identified in Escherichia coli was evaluated with respect to susceptibility to β-lactam/β-lactamase inhibitor combinations including ceftazidime-avibactam, imipenem-relebactam, meropenem-vaborbactam, aztreonam-avibactam, cefepime-taniborbactam, and to cefiderocol. A large series of E. coli recombinant strains producing broad-spectrum β-lactamases was evaluated. While imipenem-relebactam showed a similar activity regardless of the PBP3 background, susceptibility to other molecules tested was affected at various levels. This was particularly the case for ceftazidime-avibactam, aztreonam-avibactam, and cefepime-taniborbactam.

Meropenem-ANT3310, a unique β-lactam-β-lactamase inhibitor combination with expanded antibacterial spectrum against Gram-negative pathogens including carbapenem-resistant Acinetobacter baumannii.

ANT3310 is a novel broad-spectrum diazabicyclooctane serine β-lactamase inhibitor being developed in combination with meropenem (MEM) for the treatment of serious infections in hospitalized patients where carbapenem-resistant Gram-negative pathogens are expected. In this study, we evaluated the in vitro antibacterial activity of MEM in the presence of ANT3310 at 8 µg/mL against global clinical isolates that included Acinetobacter baumannii (n = 905), carbapenem-resistant Enterobacterales (CRE), carrying either oxacillinase (OXA) (n = 252) or Klebsiella pneumoniae carbapenemase (KPC) (n = 180) carbapenemases, and Pseudomonas aeruginosa (n = 502). MEM was poorly active against A. baumannii, as were MEM-vaborbactam, ceftazidime-avibactam, aztreonam-avibactam, cefepime-taniborbactam, cefepime-zidebactam, and imipenem-relebactam (MIC90 values of ≥32 µg/mL). On the other hand, MEM-ANT3310 displayed an MIC90 value of 4 µg/mL, similar to that observed with sulbactam-durlobactam, a drug developed to specifically treat A. baumannii infections. ANT3310 (8 µg/mL) additionally restored the activity of MEM against OXA- and KPC-producing CREs decreasing MEM MIC90 values from >32 µg/mL to 0.25 and 0.5 µg/mL, respectively. The combination of 8 µg/mL of both MEM and ANT3310 prevented growth of 97.5% of A. baumannii and 100% of OXA- and KPC-positive CREs, with ~90% of P. aeruginosa isolates also displaying MEM MICs ≤8 µg/mL. Furthermore, MEM-ANT3310 was efficacious in both thigh and lung murine infection models with OXA-23 A. baumannii. This study demonstrates the potent in vitro activity of the MEM-ANT3310 combination against both carbapenem-resistant A. baumannii and Enterobacterales clinical isolates, a key differentiator to other β-lactam/β-lactamase combinations.

Phylogenetic Characterization of Resistant Salmonella Strains in Typhoid Fever Patients in Nigeria.

Salmonella species are Enterobacteriaceae associated with typhoid fever. In this study, the distribution of broad-spectrum β-lactamase regulatory genes and genetic relatedness of isolates was determined. Stool samples (400) were collected from patients with fever in Dalhatu Araf Specialist Hospital (DASH), Lafia, Nigeria, between March 2020 and April 2021. Salmonella species were isolated and extended-spectrum β-lactamase distribution was determined among resistant isolates using polymerase chain reaction (PCR). Genetic relatedness of Salmonella species resistant to the 10 first-line antibiotics administered was determined among S typhi isolated. Of the 60 isolates that were confirmed to belong to the genus Salmonella, 12 (20.0%) isolates with bla SHV genes were the most prevalent, blaOXA-1 and blaCTX-M-9 were present in 5 isolates each, while blaCTX-M-4 and blaTEM genes with a prevalence of 1.7% each were the least obtained in the isolates. Two isolates had a multidrug-resistant index (MDRI) of 1, and 2 others were positive with the S typhi staG gene. Sequencing to determine their diversity showed that isolates ST36 and ST138, respectively, had MDRI = 1 and are clustered in a group with a similarity coefficient of 0.00634. The 2 isolates had the highest genetic similarity, which indicates that the genetic diversity between the isolates is low, while Salmonella strain ST313L2 had a high level of genetic distance from the other isolates. The most resistant isolates are closely related which calls for concern.

Relative inhibitory activities of the broad-spectrum β-lactamase inhibitor taniborbactam against metallo-β-lactamases.

Taniborbactam (TAN) is a novel broad-spectrum β-lactamase inhibitor with significant activity against subclass B1 metallo-β-lactamases (MBLs). Here, we showed that TAN exhibited an overall excellent activity against B1 MBLs including most NDM- and VIM-like as well as SPM-1, GIM-1, and DIM-1 enzymes, but not against SIM-1. Noteworthy, VIM-1-like enzymes (particularly VIM-83) were less inhibited by TAN than VIM-2-like. Like NDM-9, NDM-30 (also differing from NDM-1 by a single amino acid substitution) was resistant to TAN.

Impact of revised breakpoints on the categorization of susceptibility of Enterobacterales to temocillin.

To harmonize with the EUCAST breakpoints, the French Society of Microbiology introduced a change in the inhibition diameter breakpoint (17 mm versus 20 mm previously) of temocillin. We assessed the impact of the new breakpoints on categorizing susceptibility of Enterobacterales to temocillin. This was a multicentric retrospective study including all Enterobacterales isolates routinely tested for temocillin susceptibility with the disc diffusion method between 1 January 2016 and 31 July 2022 in four centres. Categorization using the breakpoints of 20 mm (French guidelines CA-SFM/EUCAST 2020 v.1.1) and 17 mm (French guidelines CA-SFM/EUCAST 2021 v1.0 and EUCAST guidelines v11.0) was performed. Overall, 36 416 Enterobacterales isolates were included. The overall rate of temocillin resistance decreased from 11.3% to 4.7% (relative difference of 58.5%) when using the 17 mm breakpoint instead of the 20 mm breakpoint, respectively. The relative change ranged from -44.0% in Klebsiella aerogenes to -72.7% in Klebsiella oxytoca. The median inhibition diameter was 23 mm (IQR 21-25). The isolates with a diameter of 20 mm appeared overrepresented, whereas those with a diameter of 18 and 19 mm were underrepresented. We therefore reviewed the diameters between 18 and 21 mm of 273 isolates. Thirty-two (11.7%) of them categorized as susceptible at first measure were controlled resistant at second measure. The new breakpoint induced a decrease in the rate of isolates categorized as resistant to temocillin, increasing therapeutic choice including for Extended-spectrum beta-lactamase-producing Enterobacterales (ESBL-PE). We suggest the bias in measuring the inhibition diameter is probably related to the fact that temocillin is considered remarkably stable against broad-spectrum β-lactamases.

Comparison of Oral Microbial Composition and Determinants Encoding Antimicrobial Resistance in Dogs and Their Owners.

Consolidated studies on animal, human, and environmental health have become very important for understanding emerging zoonotic diseases and the spread of antimicrobial resistance (AMR). The aim of this study was to analyse the oral microbiomes of healthy dogs and their owners, including determinants of AMR. Shotgun metagenomic sequencing detected 299 bacterial species in pets and their owners, from which 70 species were carried by dogs and 229 species by humans. Results demonstrated a unique microbial composition of dogs and their owners. At an order level, Bacteroidales were the most prevalent oral microbiota of dogs with significantly lower prevalence in their owners where Actinomycetales and Lactobacillales predominated. Porphyromonas and Corynebacterium were the most prevalent genera in dogs, whereas Streptococcus and Actinomyces were in animal owners. The resistances to macrolides, tetracyclines, lincosamides and Cfx family A class broad-spectrum β-lactamase were detected in both animal and human microbiomes. Resistance determinants to amphenicols, aminoglycosides, sulphonamides, and quaternary ammonium compounds were detected exceptionally in dogs. In conclusion, the study demonstrated different bacterial composition in oral microbiomes of healthy dogs without clinical signs of periodontal disease and their owners. Due to the low numbers of the samples tested, further investigations with an increased number of samples should be performed.

Culture-Independent Multiplexed Detection of Drug-Resistant Bacteria Using Surface-Enhanced Raman Scattering.

The rapid and accurate detection of bacteria resistance to β-lactam antibiotics is critical to inform optimal treatment and prevent overprescription of potent antibiotics. Here, we present a fast, culture-independent method for the detection of extended-spectrum β-lactamases (ESBLs) using surface-enhanced Raman scattering (SERS). The method uses Raman probes that release sulfur-based Raman active molecules in the presence of β-lactamases. The released thiol molecules can be captured by gold nanoparticles, leading to amplified Raman signals. A broad-spectrum cephalosporin probe R1G and an ESBL-specific probe R3G are designed to enable duplex detection of bacteria expressing broad-spectrum β-lactamases or ESBLs with a detection limit of 103 cfu/mL in 1 h incubation. Combined with a portable Raman microscope, our culturing-free SERS assay has reduced screening time to 1.5 h without compromising sensitivity and specificity.