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PubMed2026

Emergence of cefiderocol resistance in carbapenem-resistant Escherichia coli ST167 prior to clinical use: A multifactored resistance landscape.

OBJECTIVES: Cefiderocol is a novel siderophore cephalosporin with potent activity against multidrug-resistant Gram-negative bacteria. Here, we reported the prevalence and mechanisms of cefiderocol resistance in carbapenem-resistant Escherichia coli (CREC) in China before its clinical use. METHODS: A total of 443 non-duplicate CREC isolates collected from 67 hospitals in China (2013-2021) underwent antimicrobial susceptibility testing according to CLSI guidelines. Whole-genome sequencing, transcriptomic analysis, siderophore quantification, and targeted genetic manipulation were performed to investigate the underlying resistance mechanisms. RESULTS: Among the 443 CREC isolates, 102 (23.0%) were resistant to cefiderocol, and 34 (7.6%) showed intermediate susceptibility. Multivariable logistic regression identified ST167 lineage (OR, 3.05; 95% CI, 1.12-8.29; P = 0.028), blaNDM-5 carriage (OR, 9.04; 95% CI, 2.96-27.57; P < 0.001), and cirA truncation (OR, 49.56; 95% CI, 20.33-120.79; P < 0.001) as independent factors associated with cefiderocol resistance. Among ST167 isolates, cefiderocol-resistant isolates showed increased yersiniabactin carriage and siderophore production but comparable TonB-dependent transporter expression profiles. Phylogenetic analysis revealed that cefiderocol-resistant ST167 isolates clustered into a distinct subclade enriched with resistance-associated determinants, including a recurrent FhuA P50S substitution detected in 59/64 (92.2%) resistant isolates. Functional assays showed that the P50S substitution increased cefiderocol minimum inhibitory concentration (0.032-0.125 µg/mL), particularly in an NDM-5-producing background (0.032-0.5 µg/mL). CONCLUSIONS: Cefiderocol resistance is highly prevalent among high-risk ST167 CREC isolates before the clinical introduction of cefiderocol in China, highlighting the need for continued surveillance of this epidemic lineage. Cefiderocol resistance is mediated by multiple resistance determinants, and we identify the recurrent FhuA P50S substitution as a novel contributor to reduced cefiderocol susceptibility.

Antimicrobial resistance↗

PubMed2026

From resistance genes to resistance states and enzymatic context-dependence in antimicrobial resistance.

Antimicrobial resistance is often inferred from resistance genes and susceptibility phenotypes measured under standardized conditions. We argue that for many resistance genes, resistance is better viewed as a context-dependent functional state; the same gene can produce different phenotypes depending on the local microenvironment, enzyme kinetics, antibiotic exposure, and bacterial physiology.

Journal Article↗

PubMed2025

Molecular characterization of colistin resistance in carbapenem-resistant Klebsiella pneumoniae from a tertiary hospital in China.

Colistin resistance in carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a significant global health challenge, as colistin remains the last-resort antibiotic for treating multidrug-resistant K. pneumoniae infections. This study aimed to investigate the prevalence and molecular mechanisms underlying colistin resistance in CRKP (Colr-CRKP) isolates in Henan, China, from 2021 to 2024. The minimum inhibitory concentrations of colistin for 134 K. pneumoniae isolates were determined using the broth microdilution method. Whole-genome sequencing was performed using the Illumina platform to identify carbapenemase genes and sequence types (STs). Colistin resistance mechanisms were investigated, including mutations in two-component systems (pmrA/pmrB, phoP/phoQ), inactivation of the mgrB gene, and the presence of plasmid-mediated mcr genes. Most isolates were collected from intensive care units (99/134, 73.9%), with 48.5% (59/134) of patients having no documented colistin exposure history. Notably, ST11 was the predominant sequence type among Colr-CRKP isolates (113/134, 84.3%), all of which carried blaKPC-2 as the sole carbapenemase determinant. In contrast, seven non-carbapenemase-producing isolates exhibited phenotypic resistance to carbapenems. Genomic analysis revealed inactivation or loss of the mgrB gene in 53.7% (72/134) of isolates, predominantly due to insertion mutations (54/72). Although 32.8% (44/134) of isolates carried mutations in two-component systems, these alterations did not exhibit pathway-specific clustering. Intriguingly, plasmid-mediated mcr genes were detected in only 1.5% (2/134) of cases (mcr-8.2 and mcr-1.1), while 22.4% (30/134) of colistin-resistant strains lacked identifiable resistance determinants based on current detection methods. Our findings indicate that disruption of the mgrB gene is the primary mechanism of colistin resistance in ST11 CRKP clones. The emergence of resistance in 48.5% of patients without prior colistin exposure, combined with low mcr gene prevalence (1.5%) and unexplained resistance in 22.4% of isolates, suggests complex selective pressures beyond direct antimicrobial use. These findings underscore the urgent need for strengthened antimicrobial stewardship and the development of alternative therapeutic strategies to combat this high-risk pathogen.IMPORTANCEThe global rise of colistin-resistant Klebsiella pneumoniae, particularly in carbapenem-resistant Klebsiella pneumoniae (CRKP) strains, has severely restricted treatment options for multidrug-resistant infections. Our study provides the first comprehensive molecular characterization of colistin resistance in CRKP in a large tertiary hospital in central China. We identified mgrB disruption as the predominant resistance mechanism, while plasmid-mediated mcr genes were rare. Notably, nearly half of the resistant isolates occurred in patients without prior colistin exposure, suggesting alternative selective pressures driving resistance. These findings highlight the complex dynamics of colistin resistance in CRKP and underscore the need for enhanced genomic surveillance and stewardship interventions to limit further dissemination.

Colistin↗

PubMed2025

Molecular characterization of drug-resistance genes and dynamics of multidrug-resistant Salmonella spp. in waterfowl: a pre- and post-antibiotic ban surveillance in Guangdong, China from 2013 to 2023.

BACKGROUND: Multidrug-Resistant Organism (MDRO) refers to bacteria that are Resistant to three or more types of antibiotics in clinical use. The global health threat posed by multidrug-resistant (MDR) bacterial pathogens and their cross-species transmission necessitates rigorous Surveillance. This urgency is amplified in China where antibiotic growth promoters were widely used in animal husbandry until the 2020 implementation of Announcement No. 194 launched by Ministry of Agriculture and Rural Affairs (Announcement 194), banning non-therapeutic antibiotics in feed. This study conducted a decade long investigation on the correlation between antimicrobial resistance (AMR) phenotypes and genetic determinants in 314 Salmonella isolates collected from waterfowl across Guangdong Province, China, utilizing disk diffusion (Kirby-Bauer method) and PCR-based detection of antibiotic resistance genes (ARGs). The study period covered the antibiotic policy transition in China, specifically encompassing the pre-ban (2013-2019) and post-ban (2020-2023) phases of the nationwide prohibition on growth-promoting antimicrobials in animal feed. METHODS: Antimicrobial Susceptibility profiles against 16 agents were determined via Kirby-Bauer testing, while PCR amplification targeted 20 ARGs. Statistical analyses evaluated phenotype-genotype correlations using Pearson`s chi-square test. RESULTS: Surveillance revealed escalating resistance rates annually. Highest resistance prevalence was observed against β-lactams and amphenicols (92.25%), whereas amikacin exhibited the lowest resistance rate (9.55%). MDR prevalence reached 87.23%, with the AMP-CAZ-GEN-FFC-TET resistance profile predominating (51.6% of isolates). Genetic analysis identified 3 to 16 ARGs per isolate was harboring, with blaTEM demonstrating the highest detection frequency (90.76%). Significant phenotype-genotype correlations (p < 0.05) were observed for 13 genes: blaCTX-M, blaTEM, blaOXA, aacC2, aph(3')-I, aac(3)-IV, aadA1, qnrS, qnrA, clmA, floR, sulII, tetA. Notably, significant declines in resistance to aminoglycosides (e.g., gentamicin from 71.7 to 3.5%) and florfenicol (from 81.1 to 9.6%) were observed after China's 2019 antibiotic ban policy (p < 0.001), underscoring the impact of targeted antimicrobial stewardship in avian husbandry. CONCLUSIONS: Analysis of 314 waterfowl Salmonella strains revealed severe multidrug resistance (MDR) and diverse resistance genes (DRGs), with 13 DRGs linked to resistance. China's antibiotic ban reduced targeted resistance, but MDR persists alarmingly via acquired DRGs and adaptation. Continued enforcement may lower aminoglycoside/phenicol resistance, but β-lactam resistance will likely endure, worsened by transcontinental blaCTX-M spread. Critically, plasmid co-selection threatens to amplify MDR, demanding genomic surveillance. Mitigation requires boosting policy compliance, developing non-antibiotic therapies, mapping mutations, establishing cross-species barriers, and prioritizing One Health interventions to block resistance spread.

China↗

PubMed2026

Molecular epidemiology of levofloxacin-resistant Klebsiella pneumoniae and the association of plasmid-mediated quinolone resistance genes with key biological phenotypes.

UNLABELLED: Klebsiella pneumoniae is a major opportunistic pathogen in China, yet the molecular epidemiology of quinolone resistance remains poorly characterized. This study analyzed 2,433 clinical isolates from 37 Chinese hospitals (2018-2022). The overall levofloxacin-non-susceptible (NS) rate was 53.60%, with urinary tract isolates showing higher resistance. Whole-genome sequencing identified 12 plasmid-mediated quinolone resistance (PMQR) genes. Among 1,304 NS strains, 74.54% carried at least one PMQR gene (mainly qnrS, qnrB, and aac(6')-Ib-cr), and 60.20% also had quinolone resistance-determining region (QRDR) mutations. Functional studies revealed diverse phenotypic impacts. Most PMQR genes conferred low-level resistance (minimum inhibitory concentration [MIC] = 1 mg/L), while qnrB52 and qnrB91 caused high-level resistance (MIC = 8-16 mg/L). Notably, qnrB91 reduced biofilm formation, indicating a trade-off between resistance and colonization. Growth assays showed that qnrB52, qnrB91, and qnrS1 inhibited normal growth, whereas qepA1 and qnrS1 enhanced growth under ethanol stress. Most PMQR genes (except qnrB6) attenuated bacterial adhesion. qepA1 promoted intracellular survival in macrophages, suggesting a role in chronic infection. Animal models confirmed that qnrB6, qnrB7, qnrVC6, and aac(6')-Ib-cr significantly enhanced virulence. This study is the first in China to report qnrVC6 and novel gyrA mutations (Ser83Ala/Val, Asp87Phe/His) in K. pneumoniae. It systematically reveals how PMQR genes influence infection by modulating resistance, immune evasion, and pathogenicity. These findings highlight that PMQR genes contribute not only to antibiotic resistance but also to virulence, suggesting that treatment strategies should consider specific PMQR genotypes. This research provides the largest-scale molecular epidemiological data and a theoretical basis for controlling quinolone-resistant K. pneumoniae in China. IMPORTANCE: Quinolone-resistant Klebsiella pneumoniae poses a serious threat to public health, yet the role of plasmid-mediated quinolone resistance (PMQR) genes beyond antibiotic resistance remains underexplored. In this largest-scale multicenter study in China, we analyzed 2,433 clinical isolates and discovered that PMQR genes do more than just confer drug resistance-they also influence bacterial growth, stress survival, biofilm formation, and the ability to evade or persist within host immune cells. Some PMQR genes even enhance virulence in an animal model. These findings challenge the traditional view of resistance genes as mere contributors to drug failure, revealing that they can also shape infection outcomes by altering bacterial behavior. Understanding these dual roles may guide more precise treatment strategies targeting specific PMQR genotypes.

Klebsiella pneumoniae↗

PubMed2026

Narasin used as a feed additive in conventional rearing of broilers can co-select for vancomycin-resistant Enterococcus faecium through the NarAB ionophore resistance mechanisms.

OBJECTIVES: To investigate the role of the NarAB resistance mechanism in the selection of vancomycin-resistant Enterococcus faecium (VREfm) and assess the impact of ionophore feed additives, particularly narasin, on the emergence of VREfm in broiler chickens. MATERIALS AND METHODS: Three isogenic E. faecium strains with different antimicrobial resistance determinants were created by mutagenesis and conjugation and used in a controlled animal experiment. Ross 308 broiler chickens were inoculated with either a rifampicin-resistant, a rifampicin- and vancomycin-resistant or a rifampicin-, vancomycin- and narasin-resistant strain and fed diets supplemented with selected ionophores. Bacterial populations were analysed on selective Slanetz and Bartley agar to determine the presence and selection of VREfm and other vancomycin-resistant species. Bacterial inoculation strains and isolates were whole genome sequenced for species identification and to identify genetic resistance mechanisms. RESULTS: Narasin was shown to select for VREfm in broilers, with NarAB being essential for co-selection. Intrinsically vancomycin-resistant Pediococcus acidilactici and Enterococcus gallinarum were identified as part of the broilers' vancomycin-resistant resident microbiota. Notably, among the P. acidilactici isolates that were susceptibility tested, strains resistant to both vancomycin and narasin were only found in broilers fed narasin, supporting that narasin promotes the growth of narasin-resistant populations. CONCLUSION: Narasin use in broiler feed can co-select for vancomycin-resistant bacteria, including VREfm, through the NarAB mechanism. These findings emphasize the concerns associated with the use of particular ionophores in poultry and suggest that vancomycin and narasin resistance may be more widespread in the broiler microbiota than previously recognized. Further research is needed to understand the implications for antimicrobial resistance and human health.

Animals↗

PubMed2026

Investigation of pmrCAB and mcr associated resistance in colistin-resistant A. baumannii isolates.

BACKGROUND & OBJECTIVES: Colistin is one of the last-resort antibiotics for multidrug-resistant Acinetobacter baumannii. Increasing resistance to colistin limits treatment options, particularly in intensive care units (ICUs). The aim of this study was to compare the expression levels of pmrC, pmrA, and pmrB, among colistin-resistant and colistin-susceptible clinical A. baumannii isolates, to investigate the presence of plasmid-mediated mcr-1-5 genes, and to determine clonal relationships among colistin-resistant isolates. METHODS: A total of 110 A. baumannii isolates recovered from ICU patients in 2020 were included. Colistin minimum inhibitory concentrations were determined using the broth microdilution method. Expression levels of pmrC, pmrA, and pmrB were analyzed by RT-qPCR and compared with the reference strain A. baumannii ATCC 19606. Colistin-resistant isolates (Group 1) were compared with 10 randomly selected colistin-susceptible isolates (Group 2). Detection of mcr-1-5 genes was performed by in-house multiplex PCR. Clonal relationships among resistant isolates were assessed by PFGE. RESULTS: Colistin resistance was detected in 15.45% (17/110) of isolates. The median relative expression levels of pmrC, pmrB, and pmrA in colistin-resistant isolates were 47.84-fold (IQR: 19.29-67.18), 14.72-fold (IQR: 10.13-16.68), and 8.57-fold (IQR: 5.17-12.82), respectively. In colistin-susceptible isolates, the corresponding median expression levels were 5.32-fold (IQR: 3.60-7.97), 3.29-fold (IQR: 0.85-5.95), and 3.31-fold (IQR: 2.58-6.55). Expression levels were significantly higher in colistin-resistant isolates for pmrC (p < 0.001), pmrB (p = 0.002), and pmrA (p = 0.024). None of the resistant isolates carried mcr-1-5 genes. PFGE analysis revealed 12 distinct genotypes among 17 resistant isolates. INTERPRETATION & CONCLUSIONS: Colistin-resistant A. baumannii isolates exhibited significantly higher expression levels of the pmrC, pmrA, and pmrB genes compared to colistin-susceptible isolates. Among the genes evaluated, pmrC showed the largest effect size and the strongest association with the colistin-resistant phenotype. No changes were found in the mcr-1-5 genes among the isolates studied. Further studies, including genomic and functional analyses, are needed to elucidate the underlying mechanisms of these expression changes and their contribution to colistin resistance.

Journal Article↗

PubMed2026

Meropenem-Colistin Combination Mitigates Porin-Associated Carbapenem Resistance Development in Ertapenem-Mono-Resistant Enterobacterales.

BACKGROUND: Non-carbapenemase-producing Enterobacterales with isolated ertapenem resistance (ETP-mono-R) may represent an early stage in the evolution toward broader carbapenem resistance, but whether further resistance induction occurs and its underlying mechanisms remain poorly understood. METHODS: Resistance induction was assessed in three Escherichia coli, four Klebsiella pneumoniae, and two Enterobacter cloacae isolates through serial exposure to subinhibitory concentrations of meropenem (MEM), imipenem, ceftazidime-avibactam, or colistin (COL), with antibiotic-free passaging for reversion. Resistance induction under MEM+COL was evaluated separately. Whole-genome sequencing (WGS), targeted porin-gene Sanger sequencing, and transcriptional analysis were used to characterize resistance mechanisms across induction stages. RESULTS: Subinhibitory MEM exposure rapidly selected for carbapenem resistance through porin-associated alterations in a species-specific manner. E. coli accumulated loop-region mutations in ompC, while K. pneumoniae predominantly developed disruptive mutations in ompK36, both accompanied by marked transcriptional downregulation. In contrast, E. cloacae retained wild-type porins but showed increased MEM MICs, suggesting a non-porin-mediated mechanism. Subinhibitory exposure to COL alone rapidly induced colistin resistance but was associated with decreased carbapenem MICs. Co-exposure to MEM and COL significantly delayed resistance development and reduced MIC increases (all P < 0.05). Targeted sequencing of 26 non-carbapenemase-producing K. pneumoniae isolates resistant to all carbapenems revealed widespread disruptive ompK36 alterations, including the S337P substitution identified experimentally, consistent with a shared permeability-loss pathway. CONCLUSIONS: In ETP-mono-R Enterobacterales, subinhibitory carbapenem exposure promotes carbapenem resistance, with porin-associated mechanisms predominating in E. coli and K. pneumoniae. Co-exposure to COL attenuates this process, suggesting a potential strategy to delay the emergence of carbapenem resistance.

Enterobacterales↗

PubMed2026

Development of linezolid and daptomycin resistance in vancomycin resistant Enterococcus faecium during antibiotic treatment.

The increasing incidence of vancomycin-resistant enterococci (VRE) over the past decade has reduced treatment options largely to linezolid and daptomycin. However, the emergence of resistance to both agents further complicates the management of VRE infections. While the mechanisms of linezolid resistance are relatively well understood, those underlying daptomycin resistance remain less clearly defined. In this study, we analyzed genomic changes associated with the development of linezolid and daptomycin resistance in initially susceptible isolates following treatment at a Danish university hospital. Phenotypic susceptibility testing and whole-genome sequencing were performed on eight isolates obtained from the same patient. We identified two distinct Enterococcus faecium clones with different mechanisms of linezolid resistance. Linezolid resistance was associated with a G2576T mutation in the 23S rRNA gene (ST80 clone) and the presence of the poxtA gene (ST3082 clone). The ST80 clone also developed daptomycin resistance during therapy. We found that daptomycin resistance might result from either a G173R substitution in a gene annotated as an "ABC transporter ATP-binding protein (LolD)" or a nonsense mutation (Q58*) in phosphoketolase, with both alterations potentially acting synergistically, but further studies are warranted to confirm if these mutations can confer resistance. Together with these findings, the study demonstrates that a single patient may harbor multiple E. faecium clones simultaneously, highlighting the risk of treatment failure if all clones are not accurately identified.

Daptomycin↗

PubMed2026

Mechanisms of cefiderocol resistance in carbapenem-resistant Acinetobacter baumannii: a Swiss 2023-2025 collection.

OBJECTIVES: The numbers of infections caused by carbapenem-resistant Acinetobacter baumannii (CRAB) are increasing globally and present a significant burden on healthcare systems. This study describes the CRAB isolates received at the Swiss National Reference Centre for Emerging Antibiotic Resistance (NARA) over a 3-year period, from January 2022 to December 2025, and aimed to characterize the prevalence and mechanisms of FDC resistance. METHODS: Two-hundred and thirty-four non-duplicate CRAB isolates were submitted to NARA over the study period from hospitals and laboratories across Switzerland. Susceptibility testing was performed by disk diffusion and broth microdilution, according to EUCAST methodology. Whole-genome sequencing was performed on 11 isolates. ADC alleles were cloned into vector pVRL1 and transformed into Escherichia coli Top10. RESULTS: All isolates exhibited resistance to the carbapenems, and most were resistant to cephalosporins. Most isolates harboured an acquired class D carbapenemase, most frequently OXA-23 (181/234; 77.4%). One quarter of isolates were resistant to cefiderocol (FDC), exhibiting MICs ranging from 4->32 mg/L. Whole genome sequencing analyses, performed on 11 FDC-resistant isolates, identified that FDC resistance was due a combination of mechanisms including NDM and PER-production, mutations within the iron transporters, piuA and pirA, and the overexpression of ADC variants. CONCLUSIONS: This study showed that OXA-23 was the dominant mechanism of carbapenem-resistance in CRAB in Switzerland. Almost one quarter of CRAB isolates were resistant to "last resort" antimicrobial, FDC. The mechanisms of FDC resistance identified in this study emphasise that resistance to this antimicrobial is often complex and multifactorial, requiring high-resolution methods, including WGS, to identify.

Acinetobacter baumannii↗

PubMed2026

Insights into the fate and dynamics of antibiotic resistance in multidrug-resistant Bacillus cereus during in vitro simulated gastrointestinal digestion.

Bacillus cereus, an important pathogen responsible for causing foodborne diseases worldwide, releases pore-forming enterotoxins, which target host epithelial cells, leading to osmotic lysis and ultimately manifesting as diarrheal syndrome. Moreover, some B. cereus strains carry antimicrobial resistance genes that confer multidrug resistance against a spectrum of antibiotics. Characterizing the survival traits of multidrug-resistant (MDR) B. cereus strains in the intestinal microenvironment is essential for developing targeted strategies to effectively manage diarrheal foodborne diseases caused by this pathogen. This study used whole-genome sequencing (WGS) to evaluate the pre- and post-digestion toxigenic potential, antimicrobial resistance profiles, and genetic diversity of MDR B. cereus strains isolated from food samples in Guangdong Province, China. The four B. cereus isolates investigated in this study exhibited a genetic diversity, as determined by multilocus sequence typing analysis of WGS data. All four isolates produced the diarrheal toxins Hbl, Nhe, and CytK to varying levels, indicative of their potential to cause outbreaks of foodborne diseases. Each of the four isolates exhibited resistance to more than three classes of antibiotics, fulfilling the criterion for multidrug resistance. At an initial concentration of 9 log colony-forming units (CFU)/mL, the intestinal concentration of these four isolates crossed the threshold required to induce widespread diarrhea in the general population. Under rice slurry protection, all tested isolates maintained intestinal concentration beyond the threshold when the initial concentration was increased to ≥8 log CFU/mL. Moreover, the upregulations of genes associated with acid tolerance, bile tolerance and stress response were observed in the surviving MDR B. cereus isolates. Digestion markedly altered the antibiotic resistance profiles of the MDR B. cereus isolates. In the absence of a food matrix, the MDR isolates lost their resistance to imipenem, meropenem, amoxicillin-clavulanic acid, and trimethoprim-sulfamethoxazole post-digestion and was influenced by the initial concentration of the strains. In the presence of food matrix rice slurry, the effects of digestion on the antibiotic resistance of MDR B. cereus isolates can be mitigated, enabling them to maintain their antibiotic resistance to the greatest extent. Most remarkably, after digestion, the isolates Bce055 and Bce166 exhibited newly emergent resistance to cefotetan and trimethoprim-sulfamethoxazole, respectively. Our findings clarify the fate of MDR B. cereus isolates in the gastrointestinal tract and inform the development of prevention and control strategies for foodborne diseases caused by this pathogen.

Drug Resistance, Multiple, Bacterial↗

PubMed2025

Impeding pathways of intrinsic resistance in Escherichia coli confers antibiotic sensitization and resistance proofing.

Pathways of intrinsic resistance in bacteria are promising targets for novel antibiotics and resistance breakers. Here, we used a genome-wide screen to identify single gene knockouts of Escherichia coli that were hypersusceptible to trimethoprim and chloramphenicol, two chemically diverse broad-spectrum antibiotics. Among the hits from our screen, knockouts of acrB, an efflux pump, and rfaG or lpxM, both involved in cell envelope biogenesis, were hypersensitive to multiple antimicrobials and could sensitize genetically resistant E. coli strains to antibiotics. Using experimental evolution under trimethoprim pressure, we show that high drug selection regimes drove these knockouts to extinction more frequently than wild type. Among them, ΔacrB was most compromised in its ability to evolve resistance, establishing it as a promising target for "resistance proofing." At a sub-inhibitory trimethoprim concentration, however, all three knockouts adapted to the antibiotic and consequently recovered from hypersensitivity, albeit to different extents. This recovery was driven by mutations in drug-specific resistance pathways, rather than compensatory evolution, frequently involving upregulation of the drug target. Notably, resistance-conferring mutations could by-pass defects in cell wall biosynthesis more effectively than efflux even though resistant mutations did not directly engage either pathway. Since inhibiting drug-efflux emerged as a better strategy, we tested the ability of chlorpromazine, an efflux pump inhibitor (EPI), to resistance proof E. coli against trimethoprim. While qualitatively similar in the short term, genetic and pharmacological inhibition differed dramatically on an evolutionary time scale due to evolution of resistance to the EPI. Further, adaptation to the EPI-antibiotic pair also led to multidrug adaptation. The lack of concordance between genetic and pharmacological inhibition revealed a crucial lacuna in our understanding of the mutational repertoires that facilitate adaptation to antibiotics in bacteria. We propose that while intrinsic resistance mechanisms are effective targets for antibiotic sensitization, rapid evolutionary recovery may significantly limit their utility.

Escherichia coli↗

PubMed2025

Chromosomal resistance mutations facilitate acquisition of multidrug-resistant plasmids in Escherichia coli.

Bacteria can gain multiple resistance mechanisms in a single step by the acquisition of multidrug-resistant (MDR) plasmids, but it is unclear how antibiotic selection during the acquisition of MDR plasmids affects the evolution of additional resistance mechanisms. Through conjugating separate extended-spectrum β-lactamase (ESBL)- and carbapenemase-producing MDR plasmids into plasmid-naive Escherichia coli hosts, we examine the effects of acquisition of a single plasmid or co-acquisition of multiple plasmids upon fitness costs, resistance and subsequent genomic adaptation. We show that acquisition of pOXA-48, encoding OXA-48 carbapenemase, is associated with highly variable fitness costs and levels of resistance to ertapenem in transconjugants independent of the presence of pLL35. This phenomenon was not observed during the acquisition of ESBL CTX-M-15-encoding pLL35 alone. Within a single growth cycle, transconjugants receiving pOXA-48 rapidly gained parallel mutations affecting the membrane porin OmpF, or its regulators OmpR or EnvZ. These chromosomal mutations were not compensatory for the fitness costs imposed by the plasmid, nor did they provide significant increases in resistance to carbapenems in the absence of the pOXA-48. Rather, they acted synergistically with the plasmid-encoded carbapenemase, which alone only provided marginal resistance, together providing high-level resistance to ertapenem. Such rapid evolutionary processes may play an important role in plasmid dynamics within environments with strong antibiotic selection for plasmid-encoded antimicrobial resistance genes (ARGs), particularly when these ARGs provide only marginal resistance.

Escherichia coli↗

PubMed2024

Analysis of molecular epidemiological characteristics and antimicrobial susceptibility of vancomycin-resistant and linezolid-resistant Enterococcus in China.

BACKGROUND: This study investigates the distribution and characteristics of linezolid and vancomycin susceptibilities among Enterococcus faecalis (E. faecalis) and Enterococcus faecium (E. faecium) and explores the underlying resistance mechanisms. METHODS: A total of 2842 Enterococcus clinical isolates from patients were retrospectively collected, and their clinical data were further analyzed. The minimum inhibitory concentrations (MICs) of vancomycin and linezolid were validated by broth dilution method. The resistance genes optrA, cfr, vanA, vanB and vanM were investigated using polymerase chain reaction (PCR). Housekeeping genes and resistance genes were obtianed through whole-genome sequencing (WGS). RESULTS: Of the 2842 Enterococcus isolates, 88.5% (2516) originated from urine, with E. faecium accounted for 60.1% of these. The vanA gene was identified in 27/28 vancomycin resistant Enterococcus (VRE) isolates, 4 of which carried both vanA and vanM genes. The remaining strain was vanM positive. The optrA gene was identified in all E. faecalis isolates among linezolid resistant Enterococcus (LRE). E. faecium showed a higher multiple antibiotic resistance index (MAR index) compared to E. faecalis. The multi-locus sequence typing (MLST) showed the sequence type of E. faecium mainly belongs to clonal complex (CC) 17, nearly E. faecalis isolates analyzed were differentiated into 7 characteristics of sequence types (STs), among which ST16 of CC16 were the major lineage. CONCLUSION: Urine was the primary source of VRE and LRE isolates in this study. E. faecium showed higher levels of resistance compared to E. faecalis. OptrA gene was detected in 91.6% of LRE, which could explain linezolid resistance, and van genes were detected in all vancomycin resistant Enterococcus strains, while vanA was a key resistance mechanism in VRE identified in this study.

Linezolid↗

PubMed2026

Antimicrobial resistance in Staphylococcus pseudintermedius isolated from asymptomatic and symptomatic dogs in Montevideo, Uruguay: characterization of MRSP strains and genetic determinants of resistance.

Staphylococcus pseudintermedius is a common opportunistic pathogen in dogs and an increasing concern in veterinary medicine due to rising antimicrobial resistance, particularly to methicillin. This study aimed to characterize resistance profiles and genetic mechanisms in isolates from healthy and diseased dogs in Montevideo, Uruguay. A total of 133 isolates was analyzed (83 from clinical infections and 50 from asymptomatic carriers). Antimicrobial susceptibility was assessed by disk diffusion following veterinary guidelines. Resistance genes and SCCmec types were detected by PCR. Ten representative isolates underwent whole genome sequencing. High resistance rates were observed for penicillin (81%), erythromycin (49.6%), and clindamycin (45%). Overall, 48.9% of isolates were multidrug-resistant. Phenotypic resistance to oxacillin was detected in 26% of isolates; however, 23% carried mecA gene and were therefore classified as genotypic MRSP. These isolates were more frequent among dogs with clinical infections. These strains showed higher resistance to all antimicrobials tested. SCCmec type V was the most prevalent, and greater genetic diversity was found among isolates from symptomatic dogs. Genomic analysis revealed circulating strains of unassigned sequence types (STs), a variety of resistance genes within specific lineages, the circulation of SCCmec XIV cassette carrying strains, and an Oxacillin-susceptible Methicillin-resistant Staphylococcus pseudintermedius (OS-MRSP) isolate. These findings demonstrate the clinical and epidemiological relevance of S. pseudintermedius in Uruguay and the role of asymptomatic dogs as reservoirs of resistant strains. The results emphasize the need for surveillance, prudent antimicrobial use, and integrated control strategies within a One Health framework.

Animals↗

PubMed2026

A correlation analysis between arsenic and mercury resistance and the spread of antibiotic resistance genes in bacteria isolated from a highly contaminated brownfield.

Seventy-four bacterial strains from El Terronal (Asturias, Spain), a brownfield highly contaminated with mercury and arsenic but without anthropogenic antibiotic influx, were isolated and characterized in culture. No correlation was found between resistance to the aforementioned metal(loid)s and to antibiotics and biocides as tested by agar plates and disk diffusion method, as the number of strains deemed resistant to any of the analyzed antibiotics and biocides was not significantly higher among those with the highest resistance to arsenic and/or mercury. Genome sequencing of 17 of the isolated strains revealed a great number and diversity of antibiotic resistance genes (ARGs), as well as genes related to arsenic and mercury resistance, some of which were located in mobile genetic elements (MGEs). However, most of the detected ARGs were not located in MGEs, and no genes responsible for metal(loid) and antibiotic resistance were found to share an MGE. No evidence was found that resistance to arsenic and mercury influenced the dissemination of ARGs in the studied strains, although the abundance of antibiotic resistance mechanisms related to the presence of RND-type (Resistance-Nodulation-Division) efflux pumps could potentially contribute to cross-resistance.

antibiotics↗

PubMed2025

Evolutionary engineering and molecular characterization of an antimycin A-resistant Saccharomyces cerevisiae strain: the key role of pleiotropic drug resistance (PDR1).

Antimycin A, an antifungal agent that inhibits mitochondrial respiration, provides a useful model for studying resistance mechanisms. Antifungal resistance is an escalating clinical concern with limited treatment options available. To understand the molecular mechanisms of antimycin A resistance, a genetically stable, antimycin A-resistant Saccharomyces cerevisiae strain was successfully developed for the first time through an evolutionary engineering strategy, based on long-term systematic application of gradually increasing antimycin A stress in repetitive batch cultures without prior chemical mutagenesis. Comparative whole genome resequencing analysis of the evolved strain ant905-9 revealed two missense mutations in PDR1 and PRP8 genes involved in pleiotropic drug resistance and RNA splicing, respectively. Using CRISPR/Cas9 genome editing tools, the identified mutations were introduced individually and together into the reference strain, and it was confirmed that the Pdr1p.M732R mutation alone confers antimycin A-resistance in S. cerevisiae. Comparative transcriptomic analysis of the reverse-engineered Pdr1p.M732R strain showed alterations in PDR (pleiotropic drug resistance), transmembrane transport, vesicular trafficking, and autophagy pathways. Our results highlight the potential key role of PDR1 in antifungal drug resistance. This study provides new insights into mitochondrial drug resistance and the adaptive potential of yeast under respiratory stress.

Saccharomyces cerevisiae↗

PubMed2026

Whole-genome profiling of antimicrobial resistance and virulence determinants in extensively-drug resistant Pseudomonas aeruginosa isolates causing ventilator associated pneumonia in Egypt.

Among critically ill ICU patients under prolonged mechanical ventilation, Pseudomonas aeruginosa is one of the most common cause of ventilator-associated pneumonia (VAP), with antimicrobial pressure leading to emergence of multidrug, extensively drug and pandrug-resistant (PDR) strains. In Egypt, very little genomic data exist on P. aeruginosa associated with VAP. This study aimed at characterizing the antimicrobial resistance (AMR) determinants, virulence repertoire, MGEs, and sequence types of two highly drug-resistant Pseudomonas aeruginosa isolates, including one pandrug-resistant colistin-resistant isolate and one extensively drug-resistant colistin-susceptible isolate from respiratory tract of Egyptian ICU patients suffering from VAP. The two isolates were identified conventionally and confirmed to the species level using MALDI-TOF MS. Antibiotic susceptibility was assessed using the VITEK-2 Compact system and the broth microdilution method. Genome analysis was performed using PATRIC, ResFinder, CARD, and Mobile Element Finder. For both isolates, resistance was found to all antibiotics routinely tested, however, one isolate had high-level colistin resistance (MIC > 64 µg/mL), while the other isolate was still colistin susceptible. Whole-genome sequencing identified two rare sequence types, ST2023 and ST2685, both 6.5-7.6 Mb in size with a 66% GC content. The presence of 21 MGEs in the SRR36105565 genome shows that it has high genomic flexibility, including a broader resistome than other strains, such as blaVIM-2 and OXA variants, aminoglycoside-modifying enzymes, crpP, and disinfectant-resistance markers. Both isolates retained large virulence determinants including Type III and Type VI secretion systems, alginate regulation genes, quorum-sensing networks, and siderophore biosynthesis clusters. It also represents one of the first genomic studies of VAP associated PDR P. aeruginosa from Egypt. The combination of widespread AMR with intact virulence supports the potential value of future genomic surveillance efforts and improved antimicrobial stewardship in local ICUs.

Pneumonia, Ventilator-Associated↗