An antiplasmid system drives antibiotic resistance gene integration in carbapenemase-producing Escherichia coli lineages.

Plasmids carrying antibiotic resistance genes (ARG) are the main mechanism of resistance dissemination in Enterobacterales. However, the fitness-resistance trade-off may result in their elimination. Chromosomal integration of ARGs preserves resistance advantage while relieving the selective pressure for keeping costly plasmids. In some bacterial lineages, such as carbapenemase producing sequence type ST38 Escherichia coli, most ARGs are chromosomally integrated. Here we reproduce by experimental evolution the mobilisation of the carbapenemase blaOXA-48 gene from the pOXA-48 plasmid into the chromosome. We demonstrate that this integration depends on a plasmid-induced fitness cost, a mobile genetic structure embedding the ARG and a novel antiplasmid system ApsAB actively involved in pOXA-48 destabilization. We show that ApsAB targets high and low-copy number plasmids. ApsAB combines a nuclease/helicase protein and a novel type of Argonaute-like protein. It belongs to a family of defense systems broadly distributed among bacteria, which might have a strong ecological impact on plasmid diffusion.

Combining deep learning and droplet microfluidics for rapid and label-free antimicrobial susceptibility testing of colistin.

Efficient tools for rapid antibiotic susceptibility testing (AST) are crucial for appropriate use of antibiotics, especially colistin, which is now often considered a last resort therapy with extremely drug resistant Gram-negative bacteria. Here, we developed a rapid, easy and miniaturized colistin susceptibility assay based on microfluidics, which allows for culture and high-throughput analysis of bacterial samples. Specifically, a simple microfluidic platform that can easily be operated was designed to encapsulate bacteria in nanoliter droplets and perform a fast and automated bacterial growth detection in 2 h, using standardized samples. Direct bright-field imaging of compartmentalized samples proved to be a faster and more accurate detection method as compared to fluorescence-based analysis. A deep learning powered approach was implemented for the sensitive detection of the growth of several strains in droplets. The DropDeepL AST method (Droplet and Deep learning-based method for AST) developed here allowed the determination of the colistin susceptibility profiles of 21 fast-growing Enterobacterales (E. coli and K. pneumoniae), including clinical isolates with different resistance mechanisms, showing 100 % categorical agreement with the reference broth microdilution (BMD) method performed simultaneously. Direct AST of bacteria in urine samples on chip also provided accurate results in 2 h, without the need of complex sample preparation procedures. This method can easily be implemented in clinical microbiology laboratories, and has the potential to be adapted to a variety of antibiotics, especially for last-line antibiotics to optimize treatment of patients infected with multi-drug resistant strains.

Spread of carbapenemase-producing Morganella spp from 2013 to 2021: a comparative genomic study.

BACKGROUND: Morganella spp are opportunistic pathogens involved in various infections. Intrinsic resistance to multiple antibiotics (including colistin) combined with the emergence of carbapenemase producers reduces the number of active antimicrobials. The aim of this study was to characterise genetic features related to the spread of carbapenem-resistant Morganella spp. METHODS: This comparative genomic study included extensively drug-resistant Morganella spp isolates collected between Jan 1, 2013, and March 1, 2021, by the French National Reference Center (NRC; n=68) and European antimicrobial resistance reference centres in seven European countries (n=104), as well as one isolate from Canada, two reference strains from the Pasteur Institute collection (Paris, France), and two colistin-susceptible isolates from Bicêtre Hospital (Kremlin-Bicêtre, France). The isolates were characterised by whole-genome sequencing, antimicrobial susceptibility testing, and biochemical tests. Complete genomes from GenBank (n=103) were also included for genomic analysis, including phylogeny and determination of core genomes and resistomes. Genetic distance between different species or subspecies was performed using average nucleotide identity (ANI). Intrinsic resistance mechanisms to polymyxins were investigated by combining genetic analysis with mass spectrometry on lipid A. FINDINGS: Distance analysis by ANI of 275 isolates identified three groups: Morganella psychrotolerans, Morganella morganii subspecies sibonii, and M morganii subspecies morganii, and a core genome maximum likelihood phylogenetic tree showed that the M morganii isolates can be separated into four subpopulations. On the basis of these findings and of phenotypic divergences between isolates, we propose a modified taxonomy for the Morganella genus including four species, Morganella psychrotolerans, Morganella sibonii, Morganella morganii, and a new species represented by a unique environmental isolate. We propose that M morganii include two subspecies: M morganii subspecies morganii (the most prevalent) and M morganii subspecies intermedius. This modified taxonomy was supported by a difference in intrinsic resistance to tetracycline and conservation of metabolic pathways such as trehalose assimilation, both only present in M sibonii. Carbapenemase producers were mostly identified among five high-risk clones of M morganii subspecies morganii. The most prevalent carbapenemase corresponded to NDM-1, followed by KPC-2, and OXA-48. A cefepime-zidebactam combination was the most potent antimicrobial against the 172 extensively drug-resistant Morganella spp isolates in our collection from different European countries, which includes metallo-ß-lactamase producers. Lipid A analysis showed that the intrinsic resistance to colistin was associated with the presence of L-ARA4N on lipid A. INTERPRETATION: This global characterisation of, to our knowledge, the widest collection of extensively drug-resistant Morganella spp highlights the need to clarify the taxonomy and decipher intrinsic resistance mechanisms, and paves the way for further genomic comparisons. FUNDING: None.

Role of amino acid 159 in carbapenem and temocillin hydrolysis of OXA-933, a novel OXA-48 variant.

OXA-48 has rapidly disseminated worldwide and become one of the most common carbapenemases in many countries with more than 45 variants reported with, in some cases, significant differences in their hydrolysis profiles. The R214 residue, located in the ß5-ß6 loop, is crucial for the carbapenemase activity, as it stabilizes carbapenems in the active site and maintains the shape of the active site through interactions with D159. In this study, we have characterized a novel variant of OXA-48, OXA-933 with a single D159N change. To evaluate the importance of this residue, point mutations were generated (D159A, D159G, D159K, and D159W), kinetic parameters of OXA-933, OXA-48 D159G, and OXA-48 D159K were determined and compared to those of OXA-48 and OXA-244. The blaOXA-933 gene was borne on Tn2208, a 2,696-bp composite transposon made of two IS1 elements surrounded by 9 bp target site duplications and inserted into a non-self-transmissible plasmid pOXA-933 of 7,872 bp in size. Minimal inhibitory concentration values of E. coli expressing the blaOXA-933 gene or of its point mutant derivatives were lower for carbapenems (except for D159G) as compared to those expressing the blaOXA-48 gene. Steady-state kinetic parameters revealed lower catalytic efficiencies for expanded spectrum cephalosporins and carbapenems. A detailed structural analysis confirmed the crucial role of D159 in shaping the active site of OXA-48 enzymes by interacting with R214. Our work further illustrates the remarkable propensity of OXA-48-like carbapenemases to evolve through mutations at positions outside the ß5-ß6 loop, but interacting with key residues of it.

Development of an ultrafast PCR to detect clinically relevant acquired vancomycin-resistance genes from cultured enterococci.

BACKGROUND: VRE are increasingly described worldwide. Screening of hospitalized patients at risk for VRE carriage is mandatory to control their dissemination. Here, we have developed the Bfast [VRE Panel] PCR kit, a rapid and reliable quantitative PCR assay for detection of vanA, vanB, vanD and vanM genes, from solid and liquid cultures adaptable to classical and ultrafast real-time PCR platforms. METHODS: Validation was carried out on 133 well characterized bacterial strains, including 108 enterococci of which 64 were VRE. Analytical performances were determined on the CFX96 Touch (Bio-Rad) and Chronos Dx (BforCure), an ultrafast qPCR machine. Widely used culture plates and broths for enterococci selection/growth were tested. RESULTS: All targeted van alleles (A, B, D and M) were correctly detected without cross-reactivity with other van genes (C, E, G, L and N) and no interference with the different routinely used culture media. A specificity and sensitivity of 100% and 99.7%, respectively, were determined, with limits of detection ranging from 21 to 238 cfu/reaction depending on the targets. The Bfast [VRE Panel] PCR kit worked equally well on the CFX and Chronos Dx platforms, with differences in multiplexing capacities (five and four optical channels, respectively) and in turnaround time (45 and 16 minutes, respectively). CONCLUSIONS: The Bfast [VRE Panel] PCR kit is robust, easy to use, rapid and easily implementable in clinical microbiology laboratories for ultra-rapid confirmation of the four main acquired van genes. Its features, especially on Chronos Dx, seem to be unmatched compared to other tools for screening of VRE.

Antibiotic Susceptibility Profiles of Bacterial Isolates Recovered from Abscesses in Cattle and Sheep at a Slaughterhouse in Algeria.

Abscesses represent the most prominent emerging problem in the red meat industry, leading to great economic constraints and public health hazards. Data on etiological agents present in these purulent lesions in Algeria are very scarce. The aim of this study was to identify the bacteria responsible for these abscesses and to determine their antibiotic susceptibility profiles. A total of 123 samples of abscesses from 100 slaughtered sheep and 23 slaughtered cattle were cultured in several media. A total of 114 bacterial isolates were cultured from 103 abscesses. Bacteria were identified using MALDI-TOF, and antibiotic susceptibility was determined by the disk diffusion method on Mueller-Hinton agar. A total of 73.6% (n = 84) corresponded to Enterobacterales, of which four were multidrug-resistant (MDR). These isolates, together with Staphylococcus aureus, coagulase negative Staphylococci, and seven randomly chosen susceptible Escherichia coli isolates, were further characterized using WGS. Resistome analysis of the four MDR Enterobacterales isolates revealed the presence of OXA-48 carbapenemase in two Klebsiella pneumoniae ST985 and one E. coli ST10 isolates and a CTX-M-15 ESBL in one E. coli isolate ST1706. Two coagulase-negative Staphylococci isolates were found to carry the mecA gene. WGS showed the presence of different resistance genes and virulence genes. Our study revealed 5% of MDR Enterobacterales (including ESBLs and carbapenemases) identified from abscesses, thus urging the need for abscess monitoring in slaughterhouses.

A multicentre evaluation of the NG-test DetecTool OXA-23 for the rapid detection of OXA-23 carbapenemase directly from blood cultures.

Objectives: A multicentre study evaluating NG-Test DetecTool OXA-23 for the detection of OXA-23 carbapenemase directly from positive blood cultures (PBCs). Methods: The NG-Test DetecTool OXA-23 is an immunoassay that integrates a sample preparation device. We evaluated NG-Test DetecTool OXA-23 on 189 spiked and 126 clinical PBCs. The clinical samples' standard-of-care procedure consisted of bacterial identification from the first day of positivity by MALDI-TOF MS, conventional culture and antimicrobial susceptibility testing. The immunoassay results were verified molecularly. The strains used for the spiked samples consisted of well-characterized Acinetobacter baumannii and Proteus mirabilis strains. Results: The NG-Test DetecTool OXA-23 was evaluated on 315 PBCs and revealed sensitivity of 100% (95% CI: 98.21%-100.00%) and specificity of 100% (95% CI: 96.73%-100.00%). It provided 204 true-positive results for OXA-23 in 196 bottles with carbapenem-resistant A. baumannii (CRAB) and 8 bottles with carbapenem-resistant P. mirabilis and also provided 111 true-negative results. There were no false-positive and no false-negative results. Among the 315 PBCs studied, 83 clinical blood cultures collected in the ICU of a Greek university hospital, which were tested prospectively, all yielded CRAB, and OXA-23 was correctly detected in all samples from the first day of positivity using the NG-Test DetecTool OXA-23. Conclusions: The NG-Test DetecTool OXA-23 has exhibited excellent sensitivity and specificity for OXA-23 detection in PBCs and can provide valuable information for appropriate selection of antibiotic therapy and early implementation of infection control measures.

Multicenter study to assess the use of BL-DetecTool for the detection of CTX-M-type ESBLs and carbapenemases directly from clinical specimens.

Antimicrobial resistance (AMR) is one of the major public health problems worldwide. Multiple strategies have been put in place to address this problem. One of them is the rapid detection of the mechanisms of resistance, such as extended-spectrum beta-lactamases (ESBLs) and/or carbapenemases. We conducted a multicenter study that included nine European centers for the assessment of prototypes of a novel lateral flow immunoassay-based device (BL-DetecTool) for a rapid detection of ESBL (NG-Test CTX-M-MULTI DetecTool) and/or carbapenemases (NG-Test CARBA 5 DetecTool) from Enterobacterales and Pseudomonas aeruginosa in positive urine, positive blood cultures, and rectal swabs. We performed a prospective analysis between January 2021 and June 2022, including overall 22,010 samples. Based on each hospital information, the sensitivity to detect CTX-M was 84%-100%, 90.9%-100%, and 75%-100% for urine, positive blood cultures, and enriched rectal swabs, respectively. On the other hand, the sensitivity to detect carbapenemases was 42.8%-100%, 75%-100%, and 66.6%-100% for urine, positive blood cultures, and enriched rectal swab, respectively. BL-DetecTool allows a rapid and reliable detection of ESBL and carbapenemases directly from urine, positive blood cultures, or enriched rectal swabs, being an easy technique to implement in the workflow of clinical microbiology laboratories. IMPORTANCE: The assessed rapid assay to detect CTX-M beta-lactamases and carbapenemases directly from clinical samples can favor in the rapid detection of these mechanisms of resistance and hence the administration of a more adequate antimicrobial treatment.

Multidrug-resistant pathogens contaminate river water used in irrigation in disenfranchised communities.

OBJECTIVES: The contamination of fresh surface waters poses a significant burden on human health and prosperity, especially in marginalized communities with limited resources and inadequate infrastructure. Here, we performed in-depth genomic analyses of multidrug-resistant bacteria (MDR-B) isolated from Al-Oueik river water that is used for irrigation of agricultural fields in a disenfranchised area that also hosts a makeshift Syrian refugee camp. METHODS: A composite freshwater sample was filtered. Faecal coliforms were counted and extended spectrum cephalosporins and/or ertapenem resistant bacteria were screened. Isolates were identified using MALDI-TOF-MS and analysed using whole-genome sequencing (WGS) to identify the resistome, sequence types, plasmid types, and virulence genes. RESULTS: Approximately 106 CFU/100 mL of faecal coliforms were detected in the water. Four drug-resistant Gram-negative bacteria were identified, namely Escherichia coli, Klebsiella pneumoniae, Enterobacter hormaechei, and Pseudomonas otitidis. Notably, the E. coli isolate harboured blaNDM-5 and a YRIN-inserted PBP3, representing an emerging public health challenge. The K. pneumoniae isolate carried blaSHV-187 as well as mutations in the gene encoding the OmpK37 porin. Enterobacter hormaechei and P. otitidis harboured blaACT-16 and blaPOM-1, respectively. CONCLUSION: This report provides comprehensive genomic analyses of MDR-B in irrigation water in Lebanon. Our results further support that irrigation water contaminated with faecal material can be a reservoir of important MDR-B, which can spread to adjacent agricultural fields and other water bodies, posing both public health and food safety issues. Therefore, there is an urgent need to implement effective water quality monitoring and management programs to control the proliferation of antibiotic-resistant pathogens in irrigation water in Lebanon.

Pseudomonasaeruginosa antimicrobial susceptibility profiles, resistance mechanisms and international clonal lineages: update from ESGARS-ESCMID/ISARPAE Group.

SCOPE: Pseudomonas aeruginosa, a ubiquitous opportunistic pathogen considered one of the paradigms of antimicrobial resistance, is among the main causes of hospital-acquired and chronic infections associated with significant morbidity and mortality. This growing threat results from the extraordinary capacity of P. aeruginosa to develop antimicrobial resistance through chromosomal mutations, the increasing prevalence of transferable resistance determinants (such as the carbapenemases and the extended-spectrum ß-lactamases), and the global expansion of epidemic lineages. The general objective of this initiative is to provide a comprehensive update of P. aeruginosa resistance mechanisms, especially for the extensively drug-resistant (XDR)/difficult-to-treat resistance (DTR) international high-risk epidemic lineages, and how the recently approved ß-lactams and ß-lactam/ß-lactamase inhibitor combinations may affect resistance mechanisms and the definition of susceptibility profiles. METHODS: To address this challenge, the European Study Group for Antimicrobial Resistance Surveillance (ESGARS) from the European Society of Clinical Microbiology and Infectious Diseases launched the 'Improving Surveillance of Antibiotic-Resistant Pseudomonas aeruginosa in Europe (ISARPAE)' initiative in 2022, supported by the Joint programming initiative on antimicrobial resistance network call and included a panel of over 40 researchers from 18 European Countries. Thus, a ESGARS-ISARPAE position paper was designed and the final version agreed after four rounds of revision and discussion by all panel members. QUESTIONS ADDRESSED IN THE POSITION PAPER: To provide an update on (a) the emerging resistance mechanisms to classical and novel anti-pseudomonal agents, with a particular focus on ß-lactams, (b) the susceptibility profiles associated with the most relevant ß-lactam resistance mechanisms, (c) the impact of the novel agents and resistance mechanisms on the definitions of resistance profiles, and (d) the globally expanding XDR/DTR high-risk lineages and their association with transferable resistance mechanisms. IMPLICATION: The evidence presented herein can be used for coordinated epidemiological surveillance and decision making at the European and global level.

The occurrence of the carbapenemase gene, blaNDM-5, on a transmissible IncX3 plasmid in multidrug-resistant Escherichia coli isolated from a farm dog.

OBJECTIVES: In-depth phenotypic and genomic analyses on a carbapenem-resistant Escherichia coli isolate, recovered from the faeces of a farm dog in Lebanon, focusing on its antimicrobial resistance (AMR) patterns and the underlying resistome. METHODS: E. coli strain EC-106 was identified using MALDI-TOF-MS. Analyses using Carba NP, immunochromatographic assay NG Carba5, and other antimicrobial susceptibility testing were performed. Whole-genome sequencing (WGS) using the Illumina technology and different software available at the Center of Genomic Epidemiology wwere used to predict the resistome, sequence type (ST), plasmid types, and virulence genes. RESULTS: Susceptibility testing revealed that E. coli EC-106 was multi-drug resistant, including against newer antimicrobials such as imipenem-relebactam (MIC = 16 µg/mL), meropenem-vaborbactam (MIC = 16 µg/mL), and ceftazidime-avibactam (MIC > 32 µg/mL), but remained susceptible to aztreonam (MIC = 0.12 µg/mL), aztreonam-avibactam (MIC = 0.06 µg/mL), and cefiderocol (MIC = 0.5 µg/mL). WGS analyses showed that E. coli EC-106 carried 13 acquired resistance genes associated with resistance to ß-lactams (blaNDM-5 and blaTEM-1B), aminoglycosides (aac(3)-IId, aph(3')-Ia, aadA1, and aadA2), tetracyclines (tetA), amphenicols (partial catA1), macrolides (mphA), sulphonamides (sul1 and sul3), trimethoprim (dfrA12), and quaternary ammonium compounds (partial qacE). The blaNDM-5 was located on an IncX3 plasmid. The isolate was predicted to be a human pathogen (92.9%) and belonged to ST1011. CONCLUSION: To our knowledge, this is the first report of the detection of an IncX3 plasmid carrying the blaNDM-5 gene in animals in Lebanon, highlighting the severe AMR challenges in the country. Taken together, our current and previous findings suggest that blaNDM-5 might be spreading in different hosts and genetic backgrounds across clinical and non-clinical settings.

The emergence of carbapenemase-producing Enterobacterales in hospitals: a major challenge for a debilitated healthcare system in Lebanon.

Background: Carbapenem- and extended-spectrum cephalosporin-resistant Enterobacterales (CR-E and ESCR-E, respectively) are increasingly isolated worldwide. Information about these bacteria is sporadic in Lebanon and generally relies on conventional diagnostic methods, which is detrimental for a country that is struggling with an unprecedented economic crisis and a collapsing public health system. Here, CR-E isolates from different Lebanese hospitals were characterized. Materials and methods: Non-duplicate clinical ESCR-E or CR-E isolates (N = 188) were collected from three hospitals from June 2019 to December 2020. Isolates were identified by MALDI-TOF, and their antibiotic susceptibility by Kirby-Bauer disk diffusion assay. CR-E isolates (n = 33/188) were further analyzed using Illumina-based WGS to identify resistome, MLST, and plasmid types. Additionally, the genetic relatedness of the CR-E isolates was evaluated using an Infrared Biotyper system and compared to WGS. Results: Using the Kirby-Bauer disk diffusion assay, only 90 isolates out of the 188 isolates that were collected based on their initial routine susceptibility profile by the three participating hospitals could be confirmed as ESCR-E or CR-E isolates and were included in this study. This collection comprised E. coli (n = 70; 77.8%), K. pneumoniae (n = 13; 14.4%), Enterobacter spp. (n = 6; 6.7%), and Proteus mirabilis (n = 1; 1.1%). While 57 were only ESBL producers the remaining 33 isolates (i.e., 26 E. coli, five K. pneumoniae, one E. cloacae, and one Enterobacter hormaechei) were resistant to at least one carbapenem, of which 20 were also ESBL-producers. Among the 33 CR-E, five different carbapenemase determinants were identified: blaNDM-5 (14/33), blaOXA-244 (10/33), blaOXA-48 (5/33), blaNDM-1 (3/33), and blaOXA-181 (1/33) genes. Notably, 20 CR-E isolates were also ESBL-producers. The analysis of the genetic relatedness revealed a substantial genetic diversity among CR-E isolates, suggesting evolution and transmission from various sources. Conclusion: This study highlighted the emergence and broad dissemination of blaNDM-5 and blaOXA-244 genes in Lebanese clinical settings. The weak AMR awareness in the Lebanese community and the ongoing economic and healthcare challenges have spurred self-medication practices. Our findings highlight an urgent need for transformative approaches to combat antimicrobial resistance in both community and hospital settings.

Fecal carriage of extended-spectrum ß-lactamase-producing enterobacterales from hospitals and community settings in Gaza Strip, Palestine.

BACKGROUND: The fecal carriage of extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-PE) is a major driver of the global spread of these antibiotic resistance determinants. Here we determined the rate of fecal ESBL-PE carriage in pediatric hospitals and community-serving healthcare centers serving adults and children in the Gaza Strip, Palestine. METHODS: A total of 373 fecal and rectal samples were collected from different hospitals and clinics in Gaza. The antibiotic susceptibility was determined using the disk diffusion method and interpreted according to CLSI guidelines. The bacterial isolates were tested for ESBL production using phenotypic methods (double disk synergy test and growth on selective chromogenic media). BlaCTX-M, blaSHV, and blaTEM genes were sought by PCR. RESULTS: Out of the 373 isolates tested, 138 (37%) were considered ESBL positive as revealed by phenotypic tests. The prevalence of ESBLs among hospitalized patients was 39.1% (hospital setting) whereas, among outpatients attending community healthcare centers, it was 35.1% (community setting). ESBL production among Escherichia coli, Klebsiella pneumoniae, Citrobacter freundii, Proteus mirabilis, and Klebsiella aerogenes isolates was 52.8%, 39.1%, 26.7%, 2.8%, and 2.1% respectively. Meropenem and amikacin were the most effective antibiotics against ESBL producers (68.9% and 73.6% susceptibility, respectively), while only 15.2%, 22.5%, and 24.6% remained susceptible to ceftazidime, cefotaxime, and ceftriaxone, respectively. Out of 138 phenotypically ESBL-positive isolates, 98 randomly chosen were screened for blaCTX-M, blaTEM, and blaSHV genes. The prevalence rate of blaCTX-M was 45.9%, while blaTEM and blaSHV genes were detected in 16.8% and 5.2% of CTX-M-negative isolates (corresponding mostly for K. pneumoniae isolates in the case of SHV-PCR), respectively. CONCLUSIONS: The study revealed an alarmingly high prevalence of fecal carriage of ESBL-producing Enterobacterales among hospitalized children but also in the community of the Gaza Strip. In addition, 30% of ESBL-producers were already resistant to carbapenems, the treatment of choice of infections with ESBL-producers.

Emergence and rapid dissemination of highly resistant NDM-14-producing Klebsiella pneumoniae ST147, France, 2022.

BackgroundSince 2021, an emergence of New Delhi metallo-ß-lactamase (NDM)-14-producing Klebsiella pneumoniae has been identified in France. This variant with increased carbapenemase activity was not previously detected in Enterobacterales.AimWe investigated the rapid dissemination of NDM-14 producers among patients in hospitals in France.MethodsAll NDM-14-producing non-duplicate clinical isolates identified in France until June 2022 (n = 37) were analysed by whole genome sequencing. The phylogeny of NDM-14-producers among all K. pneumoniae sequence type (ST) 147 reported in France since 2014 (n = 431) was performed. Antimicrobial susceptibility testing, conjugation experiments, clonal relationship and molecular clock analysis were performed.ResultsThe 37 NDM-14 producers recovered in France until 2022 belonged to K. pneumoniae ST147. The dissemination of NDM-14-producing K. pneumoniae was linked to a single clone, likely imported from Morocco and responsible for several outbreaks in France. The gene bla NDM-14 was harboured on a 54 kilobase non-conjugative IncFIB plasmid that shared high homology with a known bla NDM-1-carrying plasmid. Using Bayesian analysis, we estimated that the NDM-14-producing K. pneumoniae ST147 clone appeared in 2020. The evolutionary rate of this clone was estimated to 5.61 single nucleotide polymorphisms per genome per year. The NDM-14 producers were highly resistant to all antimicrobials tested except to colistin, cefiderocol (minimum inhibitory concentration 2 mg/L) and the combination of aztreonam/avibactam.ConclusionHighly resistant NDM-14 producing K. pneumoniae can rapidly spread in healthcare settings. Surveillance and thorough investigations of hospital outbreaks are critical to evaluate and limit the dissemination of this clone.

Nationwide molecular epidemiology of carbapenemase-producing Citrobacter spp. in France in 2019 and 2020.

IMPORTANCE: The emergence of carbapenemase producers in Enterobacterales mostly involves Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae complex species. However, in France, we observed the emergence and the rapid dissemination of carbapenemase in Citrobacter spp. In this study, we demonstrated that a wide variety of carbapenemases is produced by many different species of Citrobacter spp. However, we clearly identify three high-risk clones of Citrobacter freundii, ST8, ST22, and ST91 that drive the spread of carbapenemase in France. This epidemiological study paves the way of further analysis that would aim to identify the virulence factors involved in this pellicular ability of these three clones to disseminate at the hospital.

An updated patent review of metallo-ß-lactamase inhibitors (2020-2023).

INTRODUCTION: Metallo-ß-lactamases (MBLs) are enzymes produced by bacteria that confer resistance to most ß-lactam antibiotics, including carbapenems, which have the broadest spectrum of activity. This resistance mechanism poses a significant threat to public health as it drastically reduces treatment options for severe bacterial infections. Developing effective inhibitors against MBLs is crucial to restore susceptibility to ß-lactam antibiotics. AREAS COVERED: This review aims to provide an updated analysis of patents describing novel MBL inhibitors and their potential therapeutic applications that were filed between January 2020 and May 2023. EXPERT OPINION: Significant advancements were made in the development of selective MBL inhibitors with zinc-binding and zinc-chelating mechanisms of action. Dual inhibitors, targeting simultaneously both serine-ß-lactamases (SBLs) and MBLs, represent an interesting alternative approach that is increasingly pertinent for the treatment of infections involving multiple ß-lactamases from different Ambler classes. Most examples of MBL-specific inhibitors were focused on the treatment of MBL-mediated infections in Enterobacterales, where IMP-1 was a more difficult target compared with VIM-1 or NDM-1, and much less on Pseudomonas aeruginosa or Acinetobacter baumannii, which are more challenging to address.