Isolation and Characterization of Extended-Spectrum ß-Lactamase Producing Escherichia coli from Pig Farms and Slaughterhouse.

Extended-spectrum ß-lactamase (ESBL) producing Escherichia coli represents a formidable challenge in the field of microbiology and public health due to its resistance to commonly used antibiotics. These strains pose a serious threat to human and animal health, underscoring the urgency of comprehensive research and surveillance. The ongoing investigation seeks ESBL producing E. coli strains from pig farms and slaughterhouses in West Bengal and Assam, India. A total of 309 samples were collected: nasal swabs (25), rectal swabs (25) from healthy pigs, pig pen soil (45), faeces (55), slaughterhouse effluents (115), and cleaning water (44). In these samples, 154 tested positive for E. coli, indicating a 49.8% prevalence. Among 154 E. coli isolates, 23 (14.9%) produced ESBLs, sourced from pig rectal swabs (7.1%), faeces (10.7%), slaughterhouse effluents (26.1%), and cleaning water (11.7%). Significantly, 4 ESBL E. coli isolates (6.6%) exclusively emerged from pig slaughterhouse effluents, displaying imipenem-resistant properties. The majority of ESBL E. coli primarily produced CTX-M and CMY, with consistent genetic markers bla CTX-M (100%) and bla CMY (82.6%). Remarkably, 2 (8.6%) of 17 ESBL E. coli isolates from pig slaughterhouse effluents carried the genetic marker bla NDM1. These findings stress implementing thorough surveillance in pig farms and local slaughterhouses. This proactive approach is crucial to identify ESBL E. coli strains, enhancing public health protection.

Draft genome sequence of carbapenem resistant Escherichia coli ST2083 carrying NDM-5, TEM-1, and CMY-42.

The draft genome sequence of the Escherichia coli strain (CREC-9) of sequence type (ST2083), which was isolated from the urine sample of a 69-year-old male and harbors the antimicrobial resistance genes TEM-1, blaCMY-42, and blaNDM-5 encoding resistance to ß-lactams, cephalosporins, carbapenems, and fluoroquinolones and coding various virulence factors, is presented here.

Survival of infection with TEM ß-lactamase-producing Escherichia coli with Pan-ß-lactam resistance.

BACKGROUND: Antimicrobial resistance is a critical global health issue, significantly contributing to patient mortality. Recent antibiotic developments have aimed to counteract carbapenemase-producing Enterobacterales; however, the impact of their use on the emergence of antibiotic resistance is unknown. This study investigates the first case of a non-carbapenemase-producing, pan-ß-lactam-resistant Escherichia coli strain from a patient previously treated with ceftolozane-tazobactam and cefiderocol. METHODS: This study describes the clinical progression of a 39-year-old ICU patient who developed multiple infections, culminating in the isolation of a pan-ß-lactam-resistant E. coli strain (EC554). The resistance profile was characterised through MIC determination, whole-genome sequencing, the use of the ß-lactam inactivation method, RT-qPCR, efflux pump inhibition assays, outer membrane protein analysis, and blaTEM transformation. FINDINGS: The EC554 isolate displayed resistance to all tested ß-lactams and ß-lactam-ß-lactamase inhibitor combinations. Whole-genome sequencing revealed four plasmids in EC554, with the only ß-lactamase gene being blaTEM-252 on the pEC554-PBR-X1-X1 plasmid. We found that the extremely resistant phenotype was attributable to a combination of different mechanisms: a high expression of TEM-252, efflux pump activity, porin loss, and PBP3 mutations. INTERPRETATION: The findings illustrate the complex interplay of multiple resistance mechanisms in E. coli, highlighting the potential for high-level resistance even without carbapenemase production. This study underscores the importance of comprehensively characterising resistance mechanisms in order to inform effective treatment strategies and mitigate the spread of resistant strains.

Long-Term Clinical and Ecological Impact of an Antimicrobial Stewardship Program on the Incidence of Carbapenem-Resistant Klebsiella pneumoniae Infections in a High-Endemic Hospital.

Carbapenem-resistant Klebsiella pneumoniae (CR-Kp) is currently a serious global concern. Antimicrobial stewardship programs (ASPs) are one of the key strategies to overcome this resistance. However, evidence about the long-term clinical and ecological impacts of ASPs is scarce. A multidisciplinary team conducted a multifaceted intervention in a CR-Kp endemic hospital over a 6-year period. We assessed the monthly long-term impacts of ASPs on carbapenem use, incidence density (ID), and crude death rates of hospital-acquired CR-Kp infections. Other variables potentially related to CR-Kp incidence and healthcare activity indicators were monitored. Carbapenem use showed a sustained reduction over the long term, with a difference of -66.19% (95% CI -87.03 to -45.34) between the expected pre-intervention trend consumption value and that obtained six years after starting the program. The ID of CR-Kp also decreased significantly and was maintained over the long term, with a relative reduction of -88.14% (95% CI; -100.4 to -75.85) at the end of the study period. The crude death rate of CR-Kp at 14 and 28 days decreased significantly after the intervention and remained steady after six years. Infection control indicator trends remained stable. This mixed ASP contributed to reducing the high incidence of infections and mortality rates of CR-Kp, achieving a sustained ecological and clinical effect.

Predominance of blaNDM- and blaIMP-Harboring Escherichia coli Belonging to Clonal Complexes 131 and 23 in a Major University Hospital.

Background and Objectives: Carbapenem resistance is a growing global challenge for healthcare, and, therefore, monitoring its prevalence and patterns is crucial for implementing targeted interventions to mitigate its impact on patient outcomes and public health. This study aimed to determine the prevalence of carbapenem resistance among Escherichia coli (E. coli) strains in the largest tertiary care hospital of the capital territory of Pakistan and to characterize the isolates for the presence of antimicrobial resistance genes. Additionally, the most prevalent sequence types were analyzed. Materials and Methods: A total of 15,467 clinical samples were collected from November 2020 to May 2022, underwent antimicrobial susceptibility testing, and were analyzed for antimicrobial resistance genes through conventional PCR and sequence typing using MLST. Results: In carbapenem-resistant E. coli (CR-EC), 74.19% of isolates harbored the blaNDM gene, with blaNDM-1 (66.96%), blaNDM-5 (12.17%), and blaNDM-7 (20.87%) variants detected. Additionally, blaIMP was found in 25.81% and blaOXA-48 in 35.48% of isolates. The presence of blaCTX-M15 and blaTEM was identified in 83.87% and 73.55% of CR-EC isolates, respectively, while armA and rmtB were detected in 40% and 65.16% of isolates, respectively. Colistin and tigecycline were the most effective drugs against CR-EC isolates, with both showing an MIC50 of 0.5 µg/mL. The MIC90 for colistin was 1 µg/mL, while for tigecycline, it was 2 µg/mL. MLST analysis revealed that the CR-EC isolates belonged to ST131 (24.52%), ST2279 (23.87%), ST3499 (16.13%), ST8051 (15.48%), ST8900 (9.68%), ST3329 (7.10%), ST88 (1.94%), and ST6293 (1.29%). The ST131 complex (70.97%) was the most prevalent, harboring 95.65% of the blaNDM gene, while the ST23 complex (18.06%) harbored 62.50% of the blaIMP gene. Conclusions: Implementing large-scale surveillance studies to monitor the spread of specific pathogens, along with active infection control policies, is crucial for the effective containment and prevention of future epidemics.

First report of a carbapenem-resistant Aeromonas veronii environmental isolate in the United States co-harboring two carbapenemase genes.

OBJECTIVES: The spread of carbapenem-resistant bacteria (CRB), and especially carbapenemase-producing CRB, is a global public health threat. Among them, Aeromonas species are of increasing concern because these emerging opportunistic pathogens are widespread in the environment and have increasingly been found to be resistant to carbapenems. The aim of this study was to investigate the genome and carbapenem-resistance determinants of Aeromonas veronii SS-M2-3, a highly carbapenem-resistant, carbapenemase-producing, river isolate from California (U.S.). METHODS: We first used disk diffusion assays to characterize the susceptibility profile to carbapenems and other antibiotics of A. veronii SS-M2-3. We next used whole-genome sequencing using the Illumina platform and bioinformatics analysis to characterize the resistome of this isolate and identify its carbapenemase genes. RESULTS: A. veronii SS-M2-3 was resistant to all carbapenems tested and amoxicillin-clavulanic acid, whereas it was sensitive to cefotaxime and all non-ß-lactam antibiotics tested. Whole genome sequencing of this isolate revealed a complex resistome that included multidrug efflux pump genes and three chromosomal ß-lactamase genes. These three genes encoded for highly conserved variants (82% to 97% amino acid identity) of the ChpA3 subclass B2 metallo-carbapenemase, OXA-12 class D carbapenemase and the FOX-2 class C ß-lactamase. This is the first report of an environmental A. veronni isolate from the U.S. co-harbouring two carbapenemase genes. CONCLUSIONS: These findings reveal that natural aquatic environments in the U.S. represent an underappreciated reservoir of carbapenem-resistant Aeromonas veronii isolates that can carry multiple carbapenemase genes.

Diagnosis and treatment challenges in a rare Clostridium infection: A case report.

The Clostridium genus includes >180 species of Gram-positive, anaerobic, sporulating bacteria. Under certain conditions, these can cause a wide range of invasive infections in humans. Clostridium paraputrificum occurs in the commensal intestinal flora and related bacteremia typically occurs secondary to an injury to the intestinal mucosa and in the presence of predisposing conditions, such as gastrointestinal disorders, malignancies, diabetes, HIV infection or neutropenia. The current study presents the case of a 70-year-old male patient, a rural resident living in poverty, with a history of alchohol consumption and cardiovascular pathology. Several initial and subsequent diagnoses were ruled out by successive investigations (e.g., stroke, meningitis, localized tetanus). Blood cultures were eventually found positive for Clostridium paraputrificum and the patient developed septic shock despite treatment with metronidazole and penicillin G. Once switched to carbapenem, the patient progressed favorably, suggesting that carbapenem could work as a first-line antibiotic treatment for Clostridium paraputrificum infections.

The OprF porin as a potential target for the restoration of carbapenem susceptibility in Pseudomonas aeruginosa expressing acquired carbapenemases.

Carbapenem resistance in Pseudomonas aeruginosa is primarily due to the acquisition of carbapenemases and is often associated with a diminution of the membrane permeability. The outer membrane protein, OprD, is a well-known route, by which carbapenems, predominantly imipenem, can enter the cell, and its loss has been associated with reduced susceptibility to imipenem. In this study, we investigated the antibiotic susceptibility patterns of isogenic P. aeruginosa mutants containing various acquired ß-lactamases, including carbapenemases, in a porin-depleted background. We identified that the deletion of oprF was associated with some recovery of susceptibility to carbapenems.

Ginkgolic Acid as a carbapenem synergist against KPC-2 positive Klebsiella pneumoniae.

The successful evolution of KPC-2 in bacteria has limited the clinical practice of carbapenems. This dilemma deteriorated the prognosis of associated infections and hence attracted increasing attention from researchers to explore alternative therapeutic options. Here, the enzyme inhibition assay was first performed to screen for a potent KPC-2 inhibitor. The synergistic effect of the candidate with carbapenems was further confirmed by checkboard minimum inhibitory concentration (MIC) assay, time-killing assay, disk diffusion method, and live/dead bacteria staining analysis. The mechanisms by which the candidate acts were subsequently explored through molecular dynamics (MD) simulations, etc. Our study found that Ginkgolic Acid (C13:0) (GA) exhibited effective KPC-2 inhibitory activity in both laboratory strain and clinical strain containing KPC-2. It could potentiate the killing effect of carbapenems on KPC-2-positive Klebsiella pnenmoniae (K. pnenmoniae). Further explorations revealed that GA could competitively bind to the active pocket of KPC-2 with meropenem (MEM) via residues Trp104, Gly235, and Leu166. The secondary structure and functional groups of KPC-2 were subsequently altered, which may be the main mechanism by which GA exerted its KPC-2 inhibitory effect. In addition, GA was also found to synergize with MEM to disrupt membrane integrity and increase membrane permeability, which may be another mechanism by which GA reinforced the bactericidal ability of carbapenems. Our study indicated that GA was a significant KPC-2 inhibitor that could prolong the lifespan of carbapenems and improve the prognosis of patients.

Characterization of acquired ß-lactamases in Pseudomonas aeruginosa and quantification of their contributions to resistance.

Pseudomonas aeruginosa is a highly problematic opportunistic pathogen that causes a range of different infections. Infections are commonly treated with ß-lactam antibiotics, including cephalosporins, monobactams, penicillins, and carbapenems, with carbapenems regarded as antibiotics of last resort. Isolates of P. aeruginosa can contain horizontally acquired bla genes encoding ß-lactamase enzymes, but the extent to which these contribute to ß-lactam resistance in this species has not been systematically quantified. The overall aim of this research was to address this knowledge gap by quantifying the frequency of ß-lactamase-encoding genes in P. aeruginosa and by determining the effects of ß-lactamases on susceptibility of P. aeruginosa to ß-lactams. Genome analysis showed that ß-lactamase-encoding genes are present in 3% of P. aeruginosa but are enriched in carbapenem-resistant isolates (35%). To determine the substrate antibiotics, 10 ß-lactamases were expressed from an integrative plasmid in the chromosome of P. aeruginosa reference strain PAO1. The ß-lactamases reduced susceptibility to a variety of clinically used antibiotics, including carbapenems (meropenem, imipenem), penicillins (ticarcillin, piperacillin), cephalosporins (ceftazidime, cefepime), and a monobactam (aztreonam). Different enzymes acted on different ß-lactams. ß-lactamases encoded by the genomes of P. aeruginosa clinical isolates had similar effects to the enzymes expressed in strain PAO1. Genome engineering was used to delete ß-lactamase-encoding genes from three carbapenem-resistant clinical isolates and increased susceptibility to substrate ß-lactams. Our findings demonstrate that acquired ß-lactamases play an important role in ß-lactam resistance in P. aeruginosa, identifying substrate antibiotics for a range of enzymes and quantifying their contributions to resistance.IMPORTANCEPseudomonas aeruginosa is an extremely problematic pathogen, with isolates that are resistant to the carbapenem class of ß-lactam antibiotics being in critical need of new therapies. Genes encoding ß-lactamase enzymes that degrade ß-lactam antibiotics can be present in P. aeruginosa, including carbapenem-resistant isolates. Here, we show that ß-lactamase genes are over-represented in carbapenem-resistant isolates, indicating their key role in resistance. We also show that different ß-lactamases alter susceptibility of P. aeruginosa to different ß-lactam antibiotics and quantify the effects of selected enzymes on ß-lactam susceptibility. This research significantly advances the understanding of the contributions of acquired ß-lactamases to antibiotic resistance, including carbapenem resistance, in P. aeruginosa and by implication in other species. It has potential to expedite development of methods that use whole genome sequencing of infecting bacteria to inform antibiotic treatment, allowing more effective use of antibiotics, and facilitate the development of new antibiotics.

Antimicrobial infusion practices within intensive care units: Carbapenem infusion preparation and administration process errors.

OBJECTIVE: To describe nurse preparation and administration of intermittent carbapenem infusions. RESEARCH METHODOLOGY/DESIGN: This observational study documented the carbapenem infusion process to adult patients in three general intensive care units. MAIN OUTCOME MEASURES: Timing and duration of infusions were observed. Volumetric analysis of infusion items was conducted to determine loss of reconstituted carbapenem during preparation and administration phases. RESULTS: Carbapenem infusions (n = 223) administered to twenty adult patients were observed. Infusion duration guidance was variable, with two ICUs following current literature recommendations, and one ICU referring to medication package insert information. Within these parameters, only 60 % of infusions complied with infusion duration. Non-compliance with planned time of administration impacted on desired dosing intervals. Incomplete delivery of intended dose was found during: sub-optimal reconstitution of vials, incorrect number of vials reconstituted, failure to administer a dose (missed dose), and discarding antibiotic residue in infusion items. Volumetric analysis of infusion items showed mean dose losses of 4.9 % and 1.2 % in discarded vials and syringes. Mean drug losses of 6.3 % and 30.8 % occurred in discarded infusion bags and infusion lines respectively. No flushing guidance or practice was observed. CONCLUSION: Incorrect nurse administration of antibiotics resulted in varying durations of infusions and the non-delivery of prescribed dose. Under-dosing has the potential to contribute to selection pressure for bacterial antibiotic resistance. The increasing frequency of intravenous delivery of antimicrobial agents through infusions requires an understanding of the required duration of administration and how to manage residual drug remaining in the intravenous line once the infusion is completed. IMPLICATIONS FOR CLINICAL PRACTICE: Flushing of administration lines is not common practice following intermittent antimicrobial infusions. Although there are multi-factorial risk factors for antimicrobial resistance in the critical care arena, nurse infusion practice must ensure that patients receive intended antimicrobial treatment. Attention must be given to the potential for antimicrobial resistance from environmental contamination with the disposal of infusion items containing undelivered antimicrobial medication.

Prior carbapenem exposure increases the incidence of ventilator-associated pneumonia in critically Ill children.

BACKGROUND: Prior antibiotic exposure has been identified as a risk factor for VAP occurrence, making it a growing concern among clinical practitioners. But there is a lack of systematic research on the types of antibiotics and the duration of exposure that influence VAP occurrence in children at current. METHODS: We retrospectively reviewed 278 children admitted to the Pediatric Intensive Care Unit (PICU) and underwent invasive mechanical ventilation (MV) between January 2020 and December 2022. Of these, 171 patients with MV duration ≥ 48 h were included in the study, with 61 of them developing VAP (VAP group) and the remaining 110 as the non-VAP group. We analyzed the relationship between early antibiotic exposure and VAP occurrence. RESULTS: The incidence of VAP was 21.94% (61/278). The VAP group had significantly longer length of hospital stay (32.00 vs. 20.00 days, p<0.001), PICU stay(25.00 vs. 10.00 days, p<0.001), and duration of mechanical ventilation(16.00 vs. 6.00 days, p<0.001) compared to the non-VAP group. The mortality in the VAP group was significantly higher than that in the non-VAP group (36.07% vs. 21.82%, p = 0.044). The VAP group had a significantly higher rate of carbapenem exposure (65.57% vs. 41.82%, p = 0.003) and duration of usage (9.00 vs. 5.00 days, p = 0.004) than the non-VAP group. Vancomycin and/or linezolid exposure rates (57.38% vs. 40.00%, p = 0.029) and duration (8 vs. 4.5 days, p = 0.010) in the VAP group were significantly higher than that in the non-VAP group, either. Multivariate logistic regression analysis identified the use of carbapenem (≥ 7 days) (OR = 5.156, 95% CI: 1.881-14.137, p = 0.001), repeated intubation (OR = 3.575, 95% CI: 1.449-8.823, p = 0.006), and tracheostomy (OR = 5.767, 95% CI:1.686-19.729, p = 0.005) as the independent risk factors for the occurrence of VAP, while early intravenous immunoglobulin (IVIG) was a protective factor against VAP (OR = 0.426, 95% CI: 0.185-0.98, p = 0.045). CONCLUSION: Prior carbapenem exposure (more than 7 days) was an independent risk factor for the occurrence of VAP. For critically ill children, reducing carbapenem use and duration as much as possible should be considered.

Piperacillin-tazobactam vs. carbapenems for treating hospitalized patients with ESBL-producing Enterobacterales bloodstream infections: A systematic review and meta-analysis.

OBJECTIVES: To meta-analyse the clinical efficacy of piperacillin-tazobactam vs. carbapenems for treating hospitalized patients affected by extended-spectrum beta-lactamase (ESBL)-producing Enterobacterales bloodstream infections (BSIs). METHODS: Two authors independently searched PubMed-MEDLINE and Scopus database up to January 17, 2024, to retrieve randomized controlled trials (RCTs) or observational studies comparing piperacillin-tazobactam vs. carbapenems for the management of hospitalized patients with ESBL-BSIs. Data were independently extracted by the two authors, and the quality of included studies was independently assessed according to ROB 2.0 or ROBINS-I tools. Mortality rate was selected as primary outcome. Meta-analysis was performed by pooling odds ratios (ORs) retrieved from studies providing adjustment for confounders using a random-effects model with the inverse variance method. RESULTS: After screening 3,418 articles, 10 studies were meta-analysed (one RCT and nine retrospective observational studies; N = 1,962). Mortality rate did not significantly differ between treatment with piperacillin-tazobactam vs. carbapenems (N = 6; OR: 1.41; 95% CI: 0.96-2.07; I² = 23.6%). The findings were consistent also in subgroup analyses assessing patients receiving empirical therapy (N = 5; OR: 1.36; 95% CI: 0.99-1.85), or patients having in ≥50% of cases urinary/biliary tract as the primary BSI source (N = 2; OR: 1.26; 95% CI: 0.84-1.89). Conversely, the mortality rate was significantly higher with piperacillin-tazobactam only among patients having in <50% of cases urinary/biliary tract as the primary source of BSI (N = 3; OR: 2.02; 95% CI: 1.00-4.07). CONCLUSIONS: This meta-analysis showed that, after performing appropriate adjustments for confounders, mortality and clinical outcome in patients having ESBL-producing Enterobacterales BSIs did not significantly differ among those receiving piperacillin-tazobactam compared to those receiving carbapenems.

Identificación de enterobacterias productoras de carbapenemasas en el Hospital Universitario Católico de Cuenca / Identification of carbapenemase producing enterobacteriaceae at the Catholic University Hospital of Cuenca / Identificação de enterobacteriaceae produtoras de carbapenemase no Hospital Universitário Católico de Cuenca

Las enterobacterias productoras de carbapenemasas desarrollan infecciones resistentes a los medicamentos en neumonía, infección del tracto urinario e infecciones relacionadas con dispositivos. Klebsiella pneumoniae, Escherichia coli y Enterobacter cloacae son amenazas de resistencia emergentes importantes a nivel mundial, lo que representa alta mortalidad y limitadas opciones de tratamiento. Objetivo: detectar la presencia de EPC de clase A, mediante la aplicación del test fenotípico de sinergia con ácido borónico en cepas de enterobacterias aisladas de superficies inertes en el Hospital Universitario Católico de Cuenca, Ecuador. Materiales y Métodos: estudio cuali-cuantitativo de tipo experimento puro de corte transversal y alcance exploratorio - descriptivo. Las enterobacterias se identificaron mediante test bioquímicos del sistema estandarizado API 20 E. Para la detección fenotípica de carbapenamasas de clase A se utilizó el método de sinergia de discos con ácido borónico y discos imipenem, meropenem y ertapenem. Resultados: se identificaron 25 géneros de enterobacterias, el 24 % fue Pseudomonas aeruginosam, el 20 % de enterobacterias fue productoras de carpapenemasas clase Am mientras que el 32 % fue resistente para los tres carbapenémicos en estudio, el 68 % mostró sensibilidad para imipenem, el 56 % para meropenem y 44 % para ertapenem. El 48 % de enterobacterias fueron resistentes a ertapenem, el 44 % a meropenem y 32 % a imipenem. Conclusiones: Enterobacterias como P. aureginosa, E. cloacae, Cronobacter spp. y E. coli presentan mecanismos de resistencia asociados a carbapenemasas clase A tipo KPC por lo que se recomienda vigilancia continua y estrategias de manejo para abordar la resistencia a carbapenémicos en entornos hospitalarios

Carbapenemase-producing Enterobacteriaceae develop drug-resistant infections in pneumonia, urinary tract infection, and device-related infections. Klebsiella pneumoniae, Escherichia coli, and Enterobacter cloacae are important emerging resistance threats globally, representing high mortality and limited treatment options. Objective: detect the presence of class A EPC, by applying the phenotypic synergy test with boronic acid in strains of enterobacteria isolated from inert surfaces at the Catholic University Hospital of Cuenca, Ecuador. Materials and Methods: Qualitative-quantitative study of pure cross-sectional experiment type and exploratorydescriptive scope. Enterobacteriaceae were identified using biochemical tests of the standardized API 20 E system. For the phenotypic detection of class A carbapenamases, the synergy method of disks with boronic acid and imipenem, meropenem and ertapenem disks was used. Results: 25 genera of enterobacteria were identified, 24 % were Pseudomonas aeruginosam, 20 % of enterobacteria were producers of class Am carbapenemases while 32 % were resistant to the three carbapenems under study, 68 % showed sensitivity to imipenem, 56 % for meropenem and 44 % for ertapenem. 48 % of enterobacteria were resistant to ertapenem, 44 % to meropenem and 32 % to imipenem. Conclusions: Enterobacteriaceae such as P. aureginosa, E. cloacae, Cronobacter spp. and E. coli present resistance mechanisms associated with class A carbapenemases type KPC, so continuous surveillance and management strategies are recommended to address resistance to carbapenems in hospital environments

Enterobacteriaceae produtoras de carbapenemases desenvolvem infecções resistentes a medicamentos em pneumonia, infecção do trato urinário e infecções relacionadas a dispositivos. Klebsiella pneumoniae, Escherichia coli e Enterobacter cloacae são importantes ameaças emergentes de resistência em todo o mundo, representando alta mortalidade e opções de tratamento limitadas. Objetivo: detectar a presença de CPE classe A, aplicando o teste de sinergia fenotípica com ácido borônico em cepas de enterobactérias isoladas de superfícies inertes no Hospital Universitário Católico de Cuenca, Equador. Materiais e Métodos: estudo cualitativo quantitativo, do tipo experimento transversal puro e escopo exploratório-descritivo. As enterobactérias foram identificadas por meio de testes bioquímicos do sistema padronizado API 20 E. Para a detecção fenotípica das carbapenamases classe A foi utilizado o método de sinergia de discos com ácido borônico e discos de imipenem, meropenem e ertapenem. Resultados: foram identificados 25 gêneros de enterobactérias, 24 % eram Pseudomonas aeruginosam, 20 % das enterobactérias eram produtoras de carbapenemases da classe Am enquanto 32 % eram resistentes aos três carbapenêmicos em estudo, 68 % apresentaram sensibilidade ao imipenem, 56 % ao meropenem e 44. % para ertapenem. 48 % das enterobactérias eram resistentes ao ertapenem, 44 % ao meropenem e 32 % ao imipenem. Conclusões: Enterobacteriaceae como P. aureginosa, E. cloacae, Cronobacter spp. e. coli apresentam mecanismos de resistência associados às carbapenemases classe A tipo KPC, portanto estratégias contínuas de vigilância e manejo são recomendadas para abordar a resistência aos carbapenêmicos em ambientes hospitalares.

Efficacy of aspergillomarasmine A/meropenem combinations with and without avibactam against bacterial strains producing multiple ß-lactamases.

The effectiveness of ß-lactam antibiotics is increasingly threatened by resistant bacteria that harbor hydrolytic ß-lactamase enzymes. Depending on the class of ß-lactamase present, ß-lactam hydrolysis can occur through one of two general molecular mechanisms. Metallo-ß-lactamases (MBLs) require active site Zn2+ ions, whereas serine-ß-lactamases (SBLs) deploy a catalytic serine residue. The result in both cases is drug inactivation via the opening of the ß-lactam warhead of the antibiotic. MBLs confer resistance to most ß-lactams and are non-susceptible to SBL inhibitors, including recently approved diazabicyclooctanes, such as avibactam; consequently, these enzymes represent a growing threat to public health. Aspergillomarasmine A (AMA), a fungal natural product, can rescue the activity of the ß-lactam antibiotic meropenem against MBL-expressing bacterial strains. However, the effectiveness of this ß-lactam/ß-lactamase inhibitor combination against bacteria producing multiple ß-lactamases remains unknown. We systematically investigated the efficacy of AMA/meropenem combination therapy with and without avibactam against 10 Escherichia coli and 10 Klebsiella pneumoniae laboratory strains tandemly expressing single MBL and SBL enzymes. Cell-based assays demonstrated that laboratory strains producing NDM-1 and KPC-2 carbapenemases were resistant to the AMA/meropenem combination but became drug-susceptible upon adding avibactam. We also probed these combinations against 30 clinical isolates expressing multiple ß-lactamases. E. coli, Enterobacter cloacae, and K. pneumoniae clinical isolates were more susceptible to AMA, avibactam, and meropenem than Pseudomonas aeruginosa and Acinetobacter baumannii isolates. Overall, the results demonstrate that a triple combination of AMA/avibactam/meropenem has potential for empirical treatment of infections caused by multiple ß-lactamase-producing bacteria, especially Enterobacterales.

Current Strategy for Targeting Metallo-ß-Lactamase with Metal-Ion-Binding Inhibitors.

Currently, antimicrobial resistance (AMR) is a serious health problem in the world, mainly because of the rapid spread of multidrug-resistant (MDR) bacteria. These include bacteria that produce ß-lactamases, which confer resistance to ß-lactams, the antibiotics with the most prescriptions in the world. Carbapenems are particularly noteworthy because they are considered the ultimate therapeutic option for MDR bacteria. However, this group of antibiotics can also be hydrolyzed by ß-lactamases, including metallo-ß-lactamases (MBLs), which have one or two zinc ions (Zn2+) on the active site and are resistant to common inhibitors of serine ß-lactamases, such as clavulanic acid, sulbactam, tazobactam, and avibactam. Therefore, the design of inhibitors against MBLs has been directed toward various compounds, with groups such as nitrogen, thiols, and metal-binding carboxylates, or compounds such as bicyclic boronates that mimic hydrolysis intermediates. Other compounds, such as dipicolinic acid and aspergillomarasmin A, have also been shown to inhibit MBLs by chelating Zn2+. In fact, recent inhibitors are based on Zn2+ chelation, which is an important factor in the mechanism of action of most MBL inhibitors. Therefore, in this review, we analyzed the current strategies for the design and mechanism of action of metal-ion-binding inhibitors that combat MDR bacteria.

[DISTRIBUTION OF CARBAPENEMASES IN TUNISIAN BURN PATIENTS]. / DISTRIBUTION GÉOGRAPHIQUE DES CARBAPÉNÉMASES CHEZ LES BRÛLÉS TUNISIENS.

Resistance to carpabenems in burns is rapidly spreading in many countries. Therefore identification of carbapenemase pathogen carriers is imperative in order to establish adequate infection control precautions and stop outbreaks of these multidrug-resistant bacteria. The aim of our study was to evaluate the distribution of carbapenemase producers in burn patients admitted to a burn center in Tunisia over 9 months. PCR for carbapenemase portage was performed in all patients within 48 hours of admission. Seventeen patients carried a single carbapenemase, 11 carried two, and 25 carried three. The enzymes detected were VIM (n=41), NDM (n=41) and OXA48 (n=32). Enzyme mapping revealed two main areas of carriage in central western Tunisia: Kairouan (NDM/OXA48) and Kasserine (NDM/VIM). Predictive factors for carriage of carbapenemase were: prior antibiotic therapy (n=24); mechanical ventilation (n=30); vascular catheterization (n=31) and a previous stay in intensive care (n=11).

Ten-year trends of antibiotics used for patients with pneumonia at long-term care hospitals in the Republic of Korea: An analysis based on national health insurance claims data.

Aim: To determine the trends in the usage of antimicrobial drugs by patients with pneumonia with prescriptions from long-term care (LTC) hospitals in the Republic of Korea. Method: This retrospective study was conducted from 2011 to 2022 using the National Health Insurance Review and Assessment Service claim data in Korea. We calculated antibiotic usage expressed as a daily defined dose (DDD) per 1000 patients per day (DID). Results: The number of patients with pneumonia in LTC hospitals increased by 2.7 times, from 30,000 in 2011 to 79,000 in 2022. Furthermore, antibiotic consumption per episode by patients with pneumonia in LTC hospitals increased from 17.14 DDD in 2011 to 18.11 DDD in 2022. Among the Access, Watch, and Reserve classification groups, the Watch group showed the highest usage; further, the Access group showed a decreasing trend, whereas the Watch and Reserve groups showed an increasing trend (p < 0.01). In the Watch group, the most commonly used antibiotic was J01CR05 (piperacillin and beta-lactamase inhibitor), followed in order by J01DD04 (ceftriaxone), J01MA12 (levofloxacin), and J01DH02 (meropenem). In the Reserve group, J01XB01 (colistin) and J01AA12 (tigecycline) were commonly used. Conclusion: The antibiotics prescribed for pneumonia in LTC hospitals have continuously increased the use of broad-spectrum antibiotics. Accordingly, appropriate use of antibiotics in LTC hospital settings and assessment of antibiotics used are warranted.

Development of an effective meropenem/KPC-2 inhibitor combination to combat infections caused by carbapenem-resistant Klebsiella pneumoniae.

The global public health threat of antibiotic resistance continues to escalate, and necessitates the implementation of urgent measures to expand the arsenal of antimicrobial drugs. This study identified a benzoxaborane compound, namely 5-chloro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (AN2178), which can inhibit the catalytic activity of the Klebsiella pneumoniae carbapenemase (KPC-2) enzyme effectively. The efficacy of AN2718 as an inhibitor for the KPC-2 enzyme was verified through various assays, including enzyme activity assays and isothermal titration calorimetry. Results of multiple biochemical assays, minimum inhibitory concentration assays and time-killing assays also showed that binding of AN2718 to KPC-2 enabled restoration of the bactericidal effect of meropenem. The survival rate of mice infected with carbapenem-resistant, high-virulence strains increased significantly upon treatment with AN2718. Most importantly, the meropenem and AN2718 combination was effective on KPC-2 mutations such as KPC-33, which evolved clinically and exhibited resistance to ceftazidime-avibactam after clinical use for a couple of years. Comprehensive safety tests both in vitro and in vivo, such as cytotoxicity, haemolytic activity and cytochrome P450 inhibition assays, demonstrated that AN2718 was safe for clinical use. These promising data indicate that AN2718 has high potential for approval for the treatment of drug resistant-bacterial infections, including those caused by ceftazidime-avibactam-resistant strains. AN2718 can be regarded as a valuable addition to the current antimicrobial armamentarium, and a promising tool to combat antimicrobial resistance.

Genomic characterisation of a multidrug-resistant Klebsiella pneumoniae co-harbouring blaNDM-1, blaKPC-2, and tet(A) isolated from the bloodstream infections of patients.

OBJECTIVES: Carbapenem-resistant Klebsiella pneumoniae (CRKP), a superbug that can be difficult or impossible to treat, has become a worldwide problem. This study presents the first report of a CRKP strain carrying a plasmid co-harbouring blaNDM-1, blaKPC-2, and tet(A) and the subsequent analysis of its genomic features. METHODS: Isolation and identification of bacteria, antimicrobial susceptibility test, whole genome sequencing, and conjugation experiments assay were conducted in clinical epidemiological investigations and plasmid genetic characterisation analysis. RESULTS: A total of 116 strains of bacteria were isolated from patients with bloodstream infections (BSI) between 2018 and 2023. A total of 89.66% of the isolates were carbapenem-resistant Enterobacteriaceae (CRE), with the majority (75/116) being CRKP. Among these, a novel plasmid co-harbouring blaNDM-1, blaKPC-2, and tet(A) simultaneously was found in CRKP46, and the three genes mediated conjugation by IS26, ISAba125, and IS26, respectively. This plasmid conferred carbapenem resistance to E. coli J53 after conjugative transfer, which was 2 times greater than that of CRKP46. CONCLUSION: The present study identified the occurrence of a rare plasmid co-harbouring blaNDM-1, blaKPC-2, and tet(A), and the spread of these genes was mediated by the corresponding mobile elements. The increased carbapenem resistance created by this novel plasmid challenges public health security and poses a potential threat to human health; therefore, it deserves attention.