Search Results (244)

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has become a major cause of bloodstream infections and poses serious challenges to clinical management because treatment options are limited. This study aimed to characterize antimicrobial resistance, virulence-associated features, and molecular epidemiology of CRKP bloodstream isolates using integrated phenotypic and genomic approaches. A total of 74 non-duplicate CRKP isolates were collected from bloodstream infections at three tertiary-care hospitals in Riyadh, Saudi Arabia, between 2022 and 2024. All isolates showed classical Klebsiella pneumoniae phenotypic characteristics, including intrinsic resistance to natural and aminopenicillins, and were classified as either multidrug-resistant (MDR) or extensively drug-resistant (XDR). Resistance to imipenem was universal, and resistance to other β-lactams and fluoroquinolones was high. Carbapenemase genes were detected in 96.0% of isolates using the GeneXpert^® Carba-R assay, with bla_OXA-48-like and bla_NDM being most common. Whole-genome sequencing demonstrated predominance of Ambler class D carbapenemases, particularly bla_OXA-232, with additional contributions from bla_NDM-1 and bla_NDM-5. Co-occurrence of carbapenemase genes was observed in a subset of isolates. Virulence analysis showed that 37.8% of isolates exhibited a hypermucoviscous phenotype, and more than half carried at least one virulence-associated determinant linked to capsule regulation or iron acquisition. In contrast, most isolates showed weak or no biofilm-forming capacity. Multilocus sequence typing revealed substantial genetic diversity but clear dominance of high-risk lineages, particularly ST147 and the emerging ST2096, both closely associated with bla_OXA-232 and bla_OXA-48-like genes. Capsular and O-antigen analysis showed a non-random distribution dominated by KL64 and O1/O2. Phylogenetic analysis was consistent with clonal expansion and suggested intra-hospital spread, with the intensive care unit serving as a key reservoir and dissemination to other wards. In conclusion, CRKP bloodstream infections in this setting are largely associated with a limited number of epidemic clones that combine extensive antimicrobial resistance with virulence-associated traits. These findings support the need for ongoing genome-based surveillance, strengthened infection control measures, and antimicrobial stewardship to limit the spread of high-risk K. pneumoniae lineages in healthcare settings. Full article

Background: Cefepime/enmetazobactam (FEP/META) is a novel fixed-dose β-lactam/β-lactamase inhibitor combination. The objective was to study the physicochemical stability of the approved daily dose in polypropylene syringes and elastomeric devices over a 24 or 72 h period to understand the feasibility of using FEP/META in prolonged infusions and its use for outpatient parenteral antibiotic therapy (OPAT). Methods: Solutions of FEP/META were prepared in 0.9% NaCl or 5% dextrose (D5W) and stored in syringes (6 g/1.5 g/48 mL) or silicone and polyisoprene elastomeric devices (EDs) at 6 g/1.5 g/120 mL and 6 g/1.5 g/240 mL: syringes were tested at 22–25 °C over a 24 h period, polyisoprene EDs at 2–8 °C over 72 h period, and silicone and polyisoprene EDs at 32 °C over a 24 h period. The solution was considered stable if it retained more than 90% of its initial concentration (Ci), no pH variation (±1 unit), no significant visual change, and with compliant subvisual examination. Liquid Chromatography–Electrospray Ionization–Quadrupole Time-of-Flight–Mass Spectrometry was utilized to identify intermediate degradation products. Results: At the daily dose, FEP/META retained >90% of its Ci up to 12 h in 0.9% NaCl and 24 h in D5W when stored in syringes. In silicone ED, stability was enhanced up to 24 h in D5W at all concentrations. The solution was chemically stable for 24 h when stored in polyisoprene ED in 0.9% NaCl at 2–8 °C. Conclusions: FEP/META combination showed prolonged stability with physicochemical integrity up to 12–24 h in all containers and conditions. It appears to be stable for prolonged infusions and for OPAT. Full article

Background/Objectives: Pseudomonas aeruginosa is an opportunistic pathogen frequently implicated in healthcare-associated infections, particularly ventilator-associated pneumonia and other device-related infections. The global emergence of carbapenem-resistant P. aeruginosa (CRPA) represents a major clinical challenge due to its limited therapeutic options and high mortality rates. Methods: Relevant clinical data were obtained from medical records. Isolates were identified via 16S PCR, and antimicrobial susceptibility testing was performed using the Vitek2 Compact system following CLSI guidelines. Carbapenemase genes (blaGES, blaKPC, blaIMP, blaNDM, blaVIM) were detected via PCR. Clonal relationships were determined via RAPD-PCR, and some sequence types were assigned according to the global P. aeruginosa MLST database. Results: In this study, 40 non-duplicate CRPA isolates were collected from 35 patients in a tertiary-care hospital in Mexico. Most isolates originated from adult patients, predominantly from tracheal aspirates (32.5%) and urine cultures (25.0%). Mechanical ventilation was the most common invasive device associated with infection, and the overall mortality rate reached 14.3%. Antimicrobial susceptibility testing showed that 95% of isolates exhibited a multidrug-resistant phenotype, with high resistance rates to ciprofloxacin (70.0%) and β-lactams. Carbapenemase genes were detected in 55% of isolates, mainly bla_IMP, bla_GES, and bla_VIM, either alone or in combination. Notably, this is the first report of ST309 (bla_IMP), ST411 (bla_GES + bla_IMP), and ST167 (bla_GES + bla_VIM) carrying carbapenemase genes in Mexico. Conclusions: These findings highlight the persistence and genetic diversity of CRPA circulating in hospital settings and emphasize the urgent need for strengthened genomic surveillance and infection control programs to prevent the spread of these high-risk multidrug-resistant clones. Full article

Methicillin-resistant Staphylococcus aureus (MRSA), a major global public health threat due to its broad resistance, urgently requires the development of new antibiotic alternatives. (‒)‒Epigallocatechin-3-gallate (EGCG) is considered a natural bioactive compound with anti-MRSA properties. The L-Lectin module of serine-rich adhesin for platelets (SraP) is considered an important target for blocking MRSA-infected hosts. This study aims to investigate the mechanism of action of EGCG against MRSA. Surface plasmon resonance (SPR), cell adhesion and invasion, biofilm formation, checkerboard assays, RNA sequencing (RNA-seq) and quantitative real-time polymerase chain reaction (qRT-PCR) were performed. The results showed that EGCG bound to SraP L Lectin with high affinity and effectively inhibited MRSA colonization. Additionally, EGCG significantly suppressed pyrimidine metabolism and downregulated related genes, thereby potentially inhibiting bacterial growth. It also markedly reduced the expression of multiple genes associated with β-lactam resistance and inhibited biofilm formation. A strong synergistic effect was observed between EGCG and the bactericidal agent ceftriaxone (CRO). When combined with 10 μg/mL EGCG, CRO required 75% less dosage and exhibited a prolonged antimicrobial effect. In conclusion, EGCG exerts anti-MRSA effects through multiple pathways and represents a promising candidate as an alternative therapeutic agent against MRSA infections. Full article

Antimicrobial resistance (AMR) continues to represent a significant global public health concern. Gram-negative bacilli (GNB) are the primary causative agents of severe nosocomial infections and possess a notable capacity to develop resistance mechanisms that restrict therapeutic options. The objective of this study was to characterize the antimicrobial susceptibility profiles of GNB isolated at a secondary-level hospital in Guadalajara, Mexico, with the aim of identifying predominant resistance patterns and the most effective therapeutic alternatives. A descriptive, retrospective, cross-sectional study was conducted using clinical isolates of Acinetobacter spp., Pseudomonas spp., Escherichia coli, Klebsiella spp., Morganella morganii, Proteus spp., and Enterobacter spp. collected during 2024. The identification and susceptibility testing were carried out using the VITEK^® 2 automated system, and the results were interpreted in accordance with CLSI guidelines. High resistance rates were observed in Acinetobacter spp. and Pseudomonas spp., particularly to carbapenems (>50% and >40%, respectively). Escherichia coli and Klebsiella spp. demonstrated resistance to third-generation cephalosporins and trimethoprim/sulfamethoxazole, exhibiting high susceptibility to amikacin and carbapenems (>90%). New-generation β-lactam/β-lactamase inhibitor combinations, such as ceftazidime/avibactam and ceftolozane/tazobactam, have demonstrated high efficacy against resistant strains. Overall, GNB isolates in this secondary-level hospital demonstrated elevated resistance levels, particularly to β-lactams and carbapenems, which pose a significant therapeutic challenge. Nevertheless, amikacin, carbapenems, and new-generation β-lactams persist as valuable therapeutic options. In order to contain the spread of multidrug-resistant organisms, it is imperative to strengthen local surveillance, optimize antibiotic stewardship, and reinforce infection control measures. Full article

Background/Objectives: Therapeutic drug monitoring (TDM) of β-lactams (BL), BL/β-lactamase inhibitor (BLI) combinations (BL/BLIc), and of fosfomycin may play a key role in optimizing antimicrobial therapy and in preventing resistance development, especially when used by continuous infusion in critically ill or immunocompromised patients. Unfortunately, analytical methods for simultaneously quantifying multiple BL/BLIc in plasma are still lacking. Methods: The aim of this study was to develop and validate two rapid, sensitive, and accurate UPLC–qTOF–MS/MS methods for the simultaneous quantification of five novel β-lactam or β-lactam/β-lactamase inhibitor combinations (ceftolozane/tazobactam, ceftazidime/avibactam, meropenem/vaborbactam, cefiderocol, and ceftobiprole) along with fosfomycin. Methods: Human plasma samples were prepared by protein precipitation using methanol containing isotopically labeled internal standards. Chromatographic separation was achieved within 10–12 min using two Agilent Poroshell columns (EC-C18 and PFP) under positive and negative electrospray ionization modes. The method was validated according to the EMA guidelines by assessing selectivity, linearity, precision, accuracy, matrix effects, extraction recovery, and stability. Results: The methods exhibited excellent linearity (R² ≥ 0.998) across the calibration ranges for all of the analytes (1.56–500 µg/mL), with limits of quantification ranging from 1.56 to 15.62 µg/mL. Intra- and inter-day precision and accuracy were always within ±15%. Extraction recovery always exceeded 92%, and the matrix effects were effectively corrected through isotopic internal standards. No carry-over or isobaric interferences were observed. All the analytes were stable for up to five days at 4 °C, but the BL and BL/BLIc stability was affected by multiple freeze–thaw cycles. Conclusions: These UPLC-qTOF-MS/MS multi-analyte methods enabled a simultaneous, reliable quantification in plasma of five novel beta-lactams and of fosfomycin. Robustness, high throughput, and sensitivity make these multi-methods feasible for real-time TDM, supporting personalized antimicrobial dosing and improved therapeutic outcomes in patients with severe or multidrug-resistant infections. Full article

Background/Objectives: Klebsiella pneumoniae is a major Gram-negative pathogen associated with community-acquired pneumonia (CAP) and a critical contributor to antimicrobial resistance (AMR). Culture-based diagnostics remain the clinical standard but may underestimate microbial diversity and resistance gene profiles. This pilot study compared pathogen detection and antimicrobial resistance gene (ARG) repertoires in matched K. pneumoniae pure cultures and primary sputum samples using targeted next-generation sequencing (tNGS). Methods: We analyzed 153 sputum samples from patients with CAP. Among 48 culture-positive cases, 22 (14% overall; 54% culture-positive) yielded K. pneumoniae. MALDI-TOF MS, phenotypic drug susceptibility testing, and tNGS were conducted on both culture isolates and matched sputum specimens. Microbial composition, ARG diversity, and method concordance were evaluated, with focused analysis of discordant and fatal cases. Results: K. pneumoniae was detected in 14.4% of all CAP cases and accounted for 54.2% of culture-positive samples. Identification rates differed across methods: 35% by MALDI-TOF MS, 45% by culture tNGS, and 29% by sputum tNGS. Sputum tNGS revealed substantially higher microbial diversity than cultures (3.04 vs. 1.42 species per sample) and detected more than sixfold unique ARGs (38 vs. 7), including clinically relevant determinants that were absent from culture isolates. Concordance was high between MALDI-TOF MS and culture tNGS (κ = 0.712), but low between sputum and culture tNGS (κ = 0.279). Among twelve K. pneumoniae isolates included in AMR analysis, all showed resistance to β-lactams, and two-thirds exhibited MDR/XDR phenotypes. Genotypic screening identified seven ARGs, but major ESBL and carbapenemase genes were not detected, suggesting the presence of alternative resistance mechanisms. Overall, sputum tNGS provided additional etiological and resistome information not captured by cultivation and complemented classical diagnostics in CAP involving K. pneumoniae. Conclusions: Culture-based diagnostics and tNGS provide complementary insights into the detection and resistance profiling of K. pneumoniae in CAP, with sputum tNGS revealing broader microbial and resistome information than pure cultures, while classical methods remain essential for species confirmation and phenotypic AST. An integrated diagnostic approach combining both methodologies may improve pathogen detection, guide antimicrobial therapy, and enhance AMR surveillance in K. pneumoniae-associated CAP. Full article

Terminalia catappa L. (Family: Combretaceae) is used globally to treat various diseases, including bacterial infections. Whilst the antibacterial activity of T. catappa has previously been tested against antibiotic-sensitive bacterial strains, the antimicrobial activity against methicillin and β-lactam-resistant pathogens has been relatively ignored. The antibacterial activity of T. catappa extracts, both alone and combined with selected clinical antibiotics, was evaluated in this study. The inhibition of bacterial growth by the extracts was determined using agar diffusion and broth micro-dilution assays. Combinations of the extracts and several clinical antibiotics were also examined and the ∑FICs were calculated to determine the interaction class. Synergistic combinations were further evaluated by isobologram analysis. The T. catappa leaf extracts were screened for toxicity using Artemia franciscana lethality bioassays (ALA). Orbitrap liquid chromatography–mass spectrometry (LC-MS) profiling analysis was undertaken to identify flavonoid components of the extracts, putatively. The T. catappa methanolic extract inhibited all the tested bacterial strains. It displayed especially good inhibitory activity against E. coli (MIC = 130 µg/mL). Combining the T. catappa extracts with some conventional antibiotics potentiated the inhibitory activity of the combinations compared to the activity of individual components. LC-MS profiling analysis identified multiple flavonoid components, including rutin, quercitin, orientin, the tannin component, and ellagic acid in the extracts. All extracts were non-toxic against Artemia nauplii. The phytochemical constituents present in the T. catappa leaf extracts warrant future investigation as potential antibacterial agents. Full article

Background/Objectives: The emergence of hypervirulent Pseudomonas aeruginosa strains resistant to β-lactamase inhibitor antibiotics is a critical health problem as they impede the treatment of infections. The objective of this study was to determine the different molecular arrangements of the virulence genotype related to β-lactamase genotype and the resistance phenotype to a combination of β-lactam antibiotics and β-lactamase inhibitors, and the phylogroups in P. aeruginosa strains isolated from patients with healthcare-associated infections and community-acquired infections. Methods: P. aeruginosa, virulence genes, β-lactamase genes and phylogroups were identified using polymerase chain reaction. Resistance to β-lactam antibiotics and β-lactamase inhibitors was determined using the disk diffusion method. The MIC determination of ticarcillin/clavulanic acid and piperacillin/tazobactam was performed using the MIC test strip for antimicrobial susceptibility testing. Results: In total, 124 P. aeruginosa strains from patients with healthcare-associated (67/124) and community-acquired infections (57/124) were analyzed. Most strains from patients with healthcare-associated infections and community-acquired infections harbored genes for proteases (aprA), phospholipases (pIcH and pIcN), elastases (lasA and lasB), rhamnolipids (rhLA), quorum-sensing system (lasI and rhII), and β-lactamase (bla_oxa-4, bla_oxa-1, and bla_GES). In total, 100% (124/124) and 99.1% (123/124) of the strains isolated from patients with healthcare-associated and community-acquired infections were resistant to the β-lactamase inhibitor antibiotics, amoxicillin/clavulanic acid and ampicillin/sulbactam, respectively, while 54% (67/124) of the strains were resistant to piperacillin/tazobactam. Phylogroup 1 (22/124) was detected more frequently among the strains in relation to phylogroup 2 (8/12). Conclusions: We demonstrated different association profiles of virulence genotype related to the β-lactamase genotype, the β-lactamase inhibitor resistome, phylogroups, and clinical origin of the strains. Therefore, medical treatment regimens against infections caused by P. aeruginosa should be improved. Full article

Background: Infections caused by the multidrug-resistant pathogen Mycobacterium abscessus (Mab) are notoriously difficult to treat. The novel β-lactamase inhibitor durlobactam, in combination with β-lactams, shows potent bactericidal activity against Mab, but the potential for acquired resistance remains a clinical concern. Objectives: To identify and characterize mechanisms of acquired resistance to durlobactam in Mab. Methods: In vitro single-step resistance selection was performed by plating wild-type Mab ATCC 19977 and by transcriptional silencing using a CRISPR interference (CRISPRi) system. Minimum inhibitory concentrations (MICs) were determined by both an agar-based method and broth microdilution. Results: Whole-genome sequencing of durlobactam-resistant mutants identified loss-of-function mutations in ponA1, a gene encoding a class A penicillin-binding protein involved in cell wall synthesis. Targeted deletion of ponA1 (ΔponA1) and CRISPRi-mediated knockdown of ponA1 expression both recapitulated the resistance phenotype, resulting in a significant increase in the durlobactam MIC on solid agar media. Strikingly, broth microdilution MICs remained largely unaffected. Conclusions: Inactivation of the peptidoglycan synthase PonA1 is a novel mechanism of resistance to durlobactam in Mab that is phenotypically expressed only during growth on solid surfaces. This finding identifies a specific genetic pathway for resistance and highlights that standard broth-based susceptibility testing could miss clinically relevant resistance mechanisms. Full article

Acinetobacter baumannii is a leading intensive care unit (ICU) pathogen associated with high rates of carbapenem resistance and poor clinical outcomes. This narrative review synthesizes recent clinical, microbiological, and pharmacokinetic/pharmacodynamic (PK/PD) evidence regarding resistance mechanisms and therapeutic strategies. A literature review was performed in PubMed, Scopus, and Web of Science (January 2015–August 2025), focusing on multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains, ICU-acquired infections, and pivotal trials involving cefiderocol and sulbactam–durlobactam. Resistance is driven by OXA-type carbapenemases (notably OXA-23/24/58), efflux systems (AdeABC/IJK/FGH), porin alterations (CarO, Omp33–36), and lipopolysaccharide (LPS) modifications conferring colistin resistance. Management options include polymyxins, optimized tigecycline dosing, β-lactam/β-lactamase inhibitors, and newer agents such as cefiderocol and sulbactam–durlobactam, though mortality and safety outcomes vary across trials. A comparative table is included, summarizing antimicrobial mechanism coverage, PK/PD parameters, and adverse effects to support regimen selection in ventilator-associated pneumonia (VAP) and bacteremia. Optimized, multimodal approaches integrating timely diagnostics, targeted combination therapies, infection prevention, and antimicrobial stewardship are essential to improve outcomes and limit the spread of MDR and XDR A. baumannii. Full article

Urinary tract infections (UTIs) are common in children and are predominantly caused by uropathogenic Escherichia coli (UPEC). An increasing proportion of these strains produce extended-spectrum β-lactamases (ESBLs), which render β-lactam antibiotics ineffective. Interestingly, some patients with ESBL-producing UTIs improve clinically following treatment with antibiotics like cephalexin, despite demonstrated in vitro resistance. Working alongside and at times synergistically with antibiotics, host immune factors, such as the antimicrobial peptide cathelicidin (LL-37), contribute to bacterial clearance through direct killing and inhibition of biofilm formation. In this review, we summarize the current understanding of pediatric ESBL-producing UPEC infections and present selected in vitro and in vivo experimental data evaluating the combined effects of LL-37 and cephalexin on clinical isolates. Although no synergy was observed, ESBL-producing isolates demonstrated reduced bacterial burden in vivo compared to a non-ESBL UPEC strain. These findings suggest that host immune factors and environmental conditions may influence the fitness and virulence of drug-resistant UTI pathogens, warranting further investigation. Full article

Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant clinical challenge due to its multidrug resistance, particularly to β-lactam antibiotics. This study comprehensively evaluated the natural compound emodin for its anti-MRSA activity, mechanisms of action, and potential for synergy with β-lactam antibiotics. Our findings demonstrate that emodin dose-dependently inhibits MRSA growth and abrogates biofilm formation at 2× MIC. Mechanistic studies revealed that emodin compromises cell membrane and wall integrity, induces oxidative stress, and downregulates the virulence factors SPA and EsxA. Furthermore, emodin acted synergistically with β-lactam antibiotics: it enhanced the ability of cefalexin to block bacterial adhesion and invasion of HaCat cells, and potentiated the efficacy of amoxicillin in clearing MRSA from infected macrophages. In conclusion, emodin employs a multi-target mechanism against MRSA and can resensitize the bacterium to conventional β-lactam antibiotics, presenting a promising strategy for combination therapy that may help curb antibiotic use and resistance development. Full article

Background/Objectives: Metallo-β-lactamase (MBL)-producing Gram-negative bacteria represent a growing global health threat due to their broad resistance to β-lactam antibiotics, including carbapenems, which severely limits treatment options. This study aimed to evaluate the in vitro synergistic activity of fosfomycin (FOS) in combination with selected older and newer antimicrobials against MBL-producing Klebsiella pneumoniae and Pseudomonas aeruginosa. Methods: Synergistic interactions were assessed using agar dilution checkerboard on 42 MBL-producing clinical isolates (22 K. pneumoniae, 20 P. aeruginosa) and confirmed using time-kill assays with selected isolates. FOS was tested in combination with colistin (COL), ceftazidime–avibactam (CAZ-AVI), meropenem (MER), amikacin (AMI), aztreonam (AZT), aztreonam–avibactam (AZT-AVI), or cefiderocol (FDC). Results: Most FOS combinations exhibited additive or synergistic effects against clinical isolates. Synergy rates reached 72.7% for the FOS+CAZ-AVI combination (K. pneumoniae) and 65.0% for the FOS+COL combination (P. aeruginosa). An asymmetric synergistic interaction was identified for FOS+CAZ-AVI, with FOS enhancing the activity of CAZ-AVI more markedly than vice versa, especially in K. pneumoniae. Time-kill assays on selected isolates confirmed synergistic and bactericidal activity of FOS+CAZ-AVI and FOS+COL, and showed that bacterial regrowth observed with FOS, CAZ-AVI, and COL alone was suppressed in combination therapy. Conclusions: FOS-based combinations, particularly with CAZ-AVI and COL, demonstrated potent synergistic activity against MBL-producing K. pneumoniae and P. aeruginosa, supporting their potential utility in rational combination therapies for infections due to MBL-producing bacteria. Full article

Urinary tract infections (UTIs) represent one of the most frequent bacterial infections worldwide, with Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) as the predominant uropathogens. The emergence of extended-spectrum β-lactamase (ESBL)-producing Enterobacterales has severely limited treatment options, making regional surveillance crucial. This study aimed to determine the prevalence of uropathogens, assess antimicrobial resistance patterns, and evaluate the burden of ESBL-producing organisms among patients presenting with suspected UTIs in a tertiary care hospital in Riyadh. We conducted a retrospective analysis of 19,556 urine cultures from a tertiary care hospital in Riyadh, Saudi Arabia, between January and December 2024. Of these, 2629 (13.4%) cultures showed significant bacterial growth, predominantly in females (83.2%) and in the 16–30 year age group. E. coli accounted for 65.9% of isolates, followed by K. pneumoniae (16.8%). ESBL production was detected in 28.5% of E. coli and Klebsiella isolates. ESBL producers exhibited complete resistance to third-generation cephalosporins and β-lactam/β-lactamase inhibitor combinations, whereas carbapenems, aminoglycosides, and fosfomycin maintained high efficacy. Resistance to ciprofloxacin and co-trimoxazole was widespread in both ESBL and non-ESBL isolates. Additionally, vancomycin-resistant enterococci (7%), methicillin-resistant Staphylococcus aureus (2%), and carbapenem-resistant Enterobacterales (0.9%) were found. These findings highlight the escalating burden of ESBL-associated UTIs and underscore the urgent need for strengthened antimicrobial stewardship, continuous surveillance, and optimized empirical therapy to mitigate the impact of multidrug-resistant uropathogens in clinical practice. Full article