Journal of Antimicrobial Chemotherapy

Two new antibiotics, zoliflodacin and gepotidacin, were recently approved for the treatment of urogenital gonorrhea. While combination therapy could, in principle, delay the emergence and spread of resistance, doing so depends on the absence of antagonism between the co-administered drugs. Using in vitro checkerboard testing, we observed no evidence of antagonism for all pairwise combinations of zoliflodacin, gepotidacin, and ciprofloxacin, including in strains with elevated ciprofloxacin MICs.

Objective: To consolidate publicly available United States National Antimicrobial Resistance Monitoring System (NARMS) human isolate data spanning 2004-2021 and comprehensively quantify trends in fluoroquinolone resistance, tetracycline resistance, ceftriaxone resistance, and early-stage azithromycin resistance across four NARMS-monitored enteric bacterial pathogens: nontyphoidal Salmonella, Campylobacter jejuni, Campylobacter coli, and Shigella spp. Methods: Annual resistance percentages were compiled from publicly available CDC, FDA, and USDA NARMS Human Isolates Surveillance Reports and Integrated Reports. Multi-year reference-period averages were assigned to their period midpoint years. Ordinary least-squares linear regression was applied to each pathogen-drug series. Slope estimates, 95% confidence intervals, Pearson correlation coefficients, and p-values were computed. A sensitivity analysis was conducted excluding reference-period midpoint years. Results: Ciprofloxacin decreased susceptibility in Salmonella rose from 2.4% to 11% (slope 0.62 pp/year, 95% CI 0.39-0.85; p less than 0.001). Ciprofloxacin resistance in C. jejuni rose from 21% to 34% (slope 1.68 pp/year, 95% CI 1.10-2.25; p=0.001) and in C. coli from 28% to 45% (slope 2.08 pp/year, 95% CI 0.97-3.19; p=0.007). Shigella showed a 5.8-fold increase (underpowered, n=3). Tetracycline resistance in C. jejuni and C. coli rose significantly. Salmonella multidrug resistance declined (slope -0.34 pp/year; p=0.001) while ceftriaxone resistance rose (slope 0.10 pp/year; p=0.028). Azithromycin-resistant Salmonella isolates detected in 2017 equalled the cumulative total from 2011-2016. Discussion: Fluoroquinolone and tetracycline resistance rose significantly across NARMS-monitored Campylobacter species, while Salmonella showed diverging trends: declining classical multidrug resistance but rising fluoroquinolone and ceftriaxone resistance. The emerging azithromycin resistance signal threatens remaining oral treatment options. These trends support sustained investment in harmonized antimicrobial resistance surveillance and stewardship globally.

Background: Most patients with melioidosis receive prolonged intravenous ceftazidime during the intensive phase of their antibiotic therapy. Contemporary guidelines use weight and renal function to guide dosing, but therapeutic drug monitoring (TDM) might enable further individualisation of therapy. Objective: To examine the potential utility of ceftazidime TDM in the management of melioidosis. Methods: We reviewed consecutive serum free ceftazidime concentrations in patients with culture-confirmed melioidosis at an Australian referral hospital. We documented the minimum inhibitory concentration (MIC) for ceftazidime Burkholderia pseudomallei isolates of the patients. We then recorded the ceftazidime dosing regimen for each patient, their serum free ceftazidime concentration and if any adverse drug reactions occurred during their treatment. Results: Trough concentrations were measured in 31 patients receiving intermittent ceftazidime dosing, while random concentrations were measured in 91 patients receiving a continuous infusion. The median (range) trough concentration:MIC ratio was 37.7 (2.7-156.6) in those receiving intermittent dosing and 47.5 (8.1-181.5) in those receiving a continuous infusion. Serum ceftazidime concentrations correlated with neurotoxicity, which was documented in 5/31 (16%) receiving intermittent dosing and in 4/91 (4%) receiving a continuous infusion. Serum ceftazidime concentrations were also higher in individuals who died from their infection than in those who survived. There was no association between ceftazidime concentrations and subsequent disease recurrence. Conclusion: Current dosing recommendations for the treatment of melioidosis achieve serum ceftazidime concentrations that greatly exceed the MIC of B. pseudomallei in this region of Australia. TDM-guided reductions in the ceftazidime dose and/or dosing frequency may mitigate the risk of ceftazidime toxicity.

Background: Tenofovir-lamivudine-dolutegravir (TLD) is an effective, single-tablet ART regimen but the causes of virologic failure that can occur despite adherence to TLD are incompletely understood, especially when resistance mutations in integrase (IN) are absent. Noncanonical resistance mutations in nucleocapsid (NC) have been selected in cell culture and are associated with decreased dolutegravir (DTG) susceptibility in vitro. Samples from the ACTG A5381/Hakim study were examined to assess the potential contribution of NC mutations to virologic failure on TLD. Methods: A5381/Hakim was an observational cohort study that enrolled individuals with HIV-1 RNA >1000 copies/mL when initiating TLD as first line ART or switching from failing non-nucleoside reverse transcriptase inhibitor (NNRTI)-based or protease inhibitor (PI)-based ART. Whole HIV RNA genome next generation sequencing was performed on paired plasma samples from study entry and confirmed virologic failure in 56 participants receiving TLD for [≥] six months. Mutations relative to the HIV-1HXB2 reference genome were identified with DeepChek(R) software using a mutation reporting threshold of five percent mixed-base frequency. Canonical drug resistance mutations (DRMs) were identified within the DeepChek(R) software using the Stanford HIVdb v9.8 algorithm. Site-directed NC and IN mutants were tested for susceptibility to DTG, raltegravir and cabotegravir in the TZM-bl HIV-1 indicator cell line and evaluated for infectivity and replication in multi-round assays. Results: Mutations in NC that emerged between TLD initiation and virologic failure were identified in 11 of 56 (20%) participants. Of these 11, three participants also had IN mutations at virologic failure but not at study entry, suggesting dual selection. Selected mutations in the NC zinc-finger domain included V13I, K20R, E21V, N27I/S, A30T, K34R, K38R, K41G/R/N, Q45R and/or M46I, alone or in combination with other NC and/or IN mutations. Specific NC mutations (K20R, N27I, A30T, K41N, M46I) conferred a significant decrease in DTG susceptibility. The combination of certain NC with IN mutations (e.g. NC N27I and IN R263K) conferred greater reduction in susceptibility to DTG in vitro than either mutation alone. Conclusions: This work provides the first clinical evidence of NC mutation selection in individuals on failing TLD ART and shows that NC mutations, combined with IN mutations, can further decrease susceptibility to DTG. Our findings support additional investigations of the contributions of mutations outside of IN to virologic failure of integrase inhibitor-containing ART regimens.

SynopsisO_ST_ABSBackgroundC_ST_ABSThe main advantages of Time Kill Curves (TKCs) in antimicrobial drug development are the ability to track bacterial kill and regrowth over time and with varying drug concentrations. Whilst there are guideline documents in place, such as M26-A in CLSI, there remains scope for individual laboratory differences in practice. Here we evaluated several factors which potentially influenced data generated in TKCs. MethodsFirstly, E. coli ATCC 25922 was used to determine optimum sampling volume, culture vessel volume, CFU enumeration variance factors and static versus agitated cultures in a single laboratory. Secondly, a ring test comprising of TKCs was performed by six laboratories focusing on: standardised inoculum, static culture and two culture vessel sizes 10 mL and 200 {micro}L. Data analysis was performed to determine consistency within centres and between them. ResultsConsistently accurate inocula could be achieved by use of: larger sampling volumes between 100 {micro}L > 20 mL; larger culture vessels volumes (10 mL > 100 {micro}L) and higher inocula (10 8 > 1.5x10 5 CFU). Culture agitation during the TKC experiment resulted in reduced killing compared to static cultures. Reproducibility of TKCs was best between centres when they were performed in 10 mL culture vessels. There was more variability per site when performing TKC in 96 well trays. ConclusionsTechnical factors such as preparation of inocula, agitation, vessel size and enumeration of cultures are important variables in performing TKCs that need to be standardised in drug development programmes involving multiple laboratory centres.

Reduced cephalosporin resistance in Enterococcus faecium has traditionally been reported in laboratory mutants and, more recently, in a single clinical ampicillin-susceptible (AmpS) isolate. Herein, we investigated whether this phenotype is widespread by analysing 95 clinical enterococcal isolates (78 AmpS and 17 ampicillin resistant [AmpR]) collected from three hospitals in Spain and Portugal (2009-2025). Low ceftriaxone MICs ([≤]4 mg/L) were detected in 19/51 (37.3%) AmpS E. faecium and 7/27 (25.9%) E. lactis but in none of the AmpR isolates. Low ceftriaxone MICs were associated with older patient age in both species and with prior ampicillin therapy in E. faecium, but not with other clinical or epidemiological variables. Ceftaroline MICs were consistently low among AmpS isolates, while ceftriaxone and cefotaxime showed greater variability. Low-MIC isolates were distributed across multiple clonal lineages and hospitals and did not share a distinctive resistance or virulence gene profile. PBP5 phylogeny and variation at the psr-pbp5 region separated AmpS from AmpR E. faecium but did not explain variability in ceftriaxone MICs. Five AmpS isolates with reduced ceftriaxone MICs carried chromosomal deletions that included the psr-pbp5 region and genes with diverse cellular functions. Variation in other candidate resistance genes (pbpA, ponA, pbpF, croRS, stpA/stk and murAA) did not consistently explain the MIC differences. These results reveal unexpected heterogeneity in intrinsic cephalosporin resistance in clinical E. faecium and E. lactis and suggest that additional genetic or regulatory mechanisms underlie reduced susceptibility.

Favipiravir (FPV) is an RdRp inhibitor developed and licensed in Japan for influenza but which has shown promising in-vitro activity against a range of RNA viruses. A physiologically-based pharmacokinetic model was developed for oral FPV and its metabolite M1 in order to optimise the dose regimen against plasma concentration targets for a number of viral pathogens. The model was validated using clinical data and was able to capture the variability in plasma concentrations for a population of individuals. FPV doses predicted to cause in-vivo exposures exceeding in-vitro IC90 targets against influenza, Ebola, Lassa fever, CCHF, SFTS, Andes virus and SARS-CoV-2, lie within the window of observed safe dosing, with SARS-CoV-2 requiring predicted doses of 2400 mg twice daily due to lower in-vitro potency. Simulations showed that a loading dose on day one of treatment should allow plasma targets to be exceeded on day one. Simulations of chronic kidney disease (CKD) showed no change to FPV plasma concentration in individuals with CKD3 and CKD5 compared to healthy individuals. Clinical data suggested active renal efflux of M1 which led to a predicted 2.2 and 11.5 fold increase in the maximum plasma concentrations of M1 in individuals with CKD3 and CKD5 respectively in comparison with healthy individuals.

SynopsisO_ST_ABSBackgroundC_ST_ABSTime kill curve (TKC) assessments are an essential step in the study of an antimicrobials pharmacodynamic characteristics. Surprisingly TKCs have not be formally standardised, therefore there remain concerns that different testing centres/methodologies may produce different results. Six centres participating in Gram-negative-Antibiotics NOW (GNA-NOW) consortium measured a series of TKCs with meropenem against E. coli to establish: Same-day (SD) vs different-day (DD) replication per centre (intra-site), and centre to centre (inter-site) correlations. MethodsMeropenem was tested against three strains of E. coli (ATCC 25922; ESBL producer C1.55; OXA-48 producer C1.62). An inoculum of 1.5x106 CFU was specified with meropenem concentrations of x0, x1 to x16 MIC; and sampling assessment of bacterial density was determined at 0-24h. Experiments were performed in triplicate, aerobically at 37{degrees}C. Centre-specific methodology was collected. Meropenem, media, bacterial strains, were shipped from one central laboratory to participating laboratories. ANOVA and Friedman tests were used to assess SD, DD and between centre replications. ResultsAssessment of the methodologies between centres revealed many differences, including bacterial inoculum, meropenem preparation, volume of TKC vessel, vessel materials, agitation vs static cultures and sampling volumes. Intra-centre SD and DD analysis for all strains were generally associated with P>0.05 suggesting consistency. Inter-centre SD and DD comparisons resulted in P<0.05, indicating variable total bacterial load measurement between centres. ConclusionsTKC methodologies varied between different centres, and while intra-centre comparison of SD and DD were generally consistent, inter-centre comparisons were not. Standardisation of TKC methodologies is required.

Background Urine tenofovir (uTFV) and dried blood spot (DBS) tenofovir diphosphate (TFV-DP) concentrations respectively estimate short- and medium-term adherence to tenofovir disoproxil fumarate (TDF)-based antiretroviral therapy (ART). We evaluated the accuracy of a point-of-care uTFV assay, and associations between uTFV/TFV-DP, and viral load (VL) and retention outcomes within a South African community ART programme. Methods We measured uTFV and DBS TFV-DP concentrations using liquid chromatography-tandem mass spectrometry (LC-MS/MS). We calculated sensitivity and specificity of the point-of-care uTFV assay at the manufacturer-recommended threshold of [&ge;]1,500 ng/mL compared to LC-MS/MS. We assessed associations of the point-of-care uTFV assay, and DBS TFV-DP concentrations with concurrent viraemia, and with retention-in-care by 16 weeks post-enrolment. Results Of 196 adults median age was 44 years, 127 (64.8%) were female, and 191 (97.4%) were receiving TDF. 185 (94.4%) had detectable point-of-care uTFV, which had high sensitivity (99.5%, 95% CI 96.5-100%) and moderate specificity (76.9%, 95% CI 46.0-93.8%) for detecting uTFV [&ge;]1,500 ng/mL. Two participants had concurrent viraemia [&ge;]1,000 copies/mL; of these 50.0% (95% CI 9.4-90.5) had undetectable point-of-care uTFV, and 100% (95% CI 19.7-100) had low TFV-DP <483 fmol/punch. Among participants without viraemia 97.4% (95% CI 93.6-99.0) had detectable uTFV, and 98.1% (95.3-99.6) had high TFV-DP [&ge;]483 fmol/punch. Point-of-care uTFV and DBS TFV-DP were not associated with retention-in-care. Conclusions The point-of-care uTFV assay demonstrated high sensitivity and moderate specificity to detect uTFV. Over 95% of people without viraemia had detectable point-of-care uTFV or high DBS TFV-DP levels respectively, but these were not associated with 16-week retention-in-care.

Resistance to {beta}-lactam antibiotics, including carbapenems, mediated by metallo-{beta}-lactamases (MBLs), including the New Delhi metallo-{beta}-lactamase (NDM) MBL subfamily, is increasing. No MBL inhibitors are currently approved for clinical use with most reported MBL inhibitors are metal ion chelators, acting either at the Zn(II) ion active site and/or in solution. The hexokinase inhibitor 3-bromopyruvate (3-BP) is reported to inhibit NDM-1. We found that 3-BP selectively restored the antimicrobial activity of meropenem against carbapenem resistant Escherichia coli, Klebsiella pneumoniae and Acinetobacter baumannii strains, obtained from clinical and environmental isolates from Tanzania and Malawi, containing genes that encode NDM-1 or NDM-5, but not against strains containing genes encoding for serine {beta}-lactamases. Mass spectrometry studies with NDM-1 and NDM-5 support a mechanism involving covalent reaction of 3-BP with an active site cysteine residue. The results will promote work on the development of covalently reacting MBL inhibitors, a strategy that has been successful for inhibition of the nucleophilic serine {beta}-lactamases.

The global emergence of multidrug-resistant (MDR) ESKAPE pathogens has significantly reduced the effectiveness of existing antibiotics, highlighting the urgent need for new strategies to restore antimicrobial susceptibility. Here, we report the discovery and mechanism of BMM_1567, a peptide potentiator that enhances aminoglycoside efficacy against MDR pathogens. A genetically defined reporter-based screen identified BMM_1567 as a potent inhibitor of aminoglycoside resistance, potentiating spectinomycin activity against MDR Gram-negative ESKAPE isolates at low micromolar concentrations. Structural modeling and molecular dynamics simulations indicated that BMM_1567 interacts with residues lining the antibiotic-binding groove of aminoglycoside-modifying enzymes (ANT, APH, AAC), with highest affinity for ANT ({Delta}G_bind = -62.25 kcal/mol), suggesting competitive inhibition of substrate binding. Site-directed mutagenesis of key ANT residues identified critical amino acids involved in BMM_1567 binding, confirming their role in mediating spectinomycin potentiation. In murine abscess model using XDR E. coli, BMM_1567 in combination with spectinomycin significantly reduced bacterial burden and pro-inflammatory cytokine levels, comparable to colistin. Collectively, these findings establish BMM_1567 as a promising aminoglycoside potentiator that restores antibiotic activity against MDR pathogens through direct inhibition of resistance enzymes, while exhibiting in vivo efficacy and a remarkably low propensity for resistance development.

Background and Aims The optimal treatment for first episodes and first recurrences of Clostridioides difficile infections (CDI) is unknown and there is emerging evidence for pulse and taper (P-T) regimens. Therefore, we sought to estimate the relative efficacy of treatment options. Methods MEDLINE and CENTRAL were searched from database inception to May 21, 2025 and unpublished conference abstracts were searched from recent infectious disease conferences. RCTs on the treatment of first episodes or first recurrences of CDI comparing fixed-dose or P-T regimens of fidaxomicin or vancomycin were included. The primary and secondary outcomes were 40- and 56-day CDI recurrence, respectively. A random-effects network meta-analysis on the risk ratio (RR) scale was conducted using a standard regimen (10-14 days) of vancomycin as the comparator. Treatments were ranked using the surface under the cumulative ranking curve (SUCRA). Results 8 RCTs were included comprising a total of 2181 patients. For 40-day recurrence, fidaxomicin P-T had the highest probability of ranking best (RR=0.10, 95%Confidence Interval [95%CI]=0.10-0.49, SUCRA=1.00), followed by vancomycin P-T (RR=0.49, 95%CI=0.32-0.76, SUCRA=0.61), fixed-dose fidaxomicin (RR=0.61, 95%CI=0.49-0.76, SUCRA=0.39), and, finally, fixed-dose of vancomycin (SUCRA=0.00). The treatments ranked in the same order for 56-day recurrence, though only 3 RCTs reported on this timepoint. Conclusion Vancomycin P-T, fidaxomicin P-T, and fixed-dose fidaxomicin were all superior to a fixed-dose vancomycin. Head-to-head comparative effectiveness RCTs are needed to quantify their relative effect sizes of and impact on long-term prevention of recurrent CDI.

The minimum inhibitory concentration (MIC) assay is the gold standard for evaluating antimicrobial activity1. However, conventional agar-based MIC methods often underestimate the potency of physicochemically complex compounds[1, 2]. Hydrophobic and adhesive molecules, such as lipoglycopeptide antibiotics, exhibit poor diffusion and non-specific binding to agar, leading to artificially elevated MIC values compared to broth-based methods[3]. This issue complicates accurate potency assessment and is particularly an issue when attempting resistance frequency (FOR) studies, which must be conducted on solid media. Here, we developed a modified miniaturised agar MIC assay using 1% agarose, 0.002% Tween 80-supplemented tryptic soy broth (TSB), and a 24-well plate format[4]. These modifications improved compound dispersion, reduced matrix interactions, and lowered compound requirements. The optimised assay was validated with vancomycin, oritavancin, and dalbavancin against Staphylococcus aureus ATCC 43300 (MRSA) and Streptococcus pneumoniae ATCC 700677. This efficient, cost-effective, high-throughput platform overcomes the limitations of traditional agar methods, enhancing reliability in evaluating challenging antimicrobials and supporting next-generation antibiotic development.

Antimalarial drug resistance remains one of the most significant challenges to global malaria control. The emergence of resistance to chloroquine, the first truly globally distributed antimalarial, has been extensively studied but is still not fully understood. While mutations in the Plasmodium falciparum chloroquine resistance transporter (PfCRT) primarily drive resistance, the phenotype is complex and multigenic, with mutations in the putative amino acid transporter PfAAT1 recently confirmed to play a modulatory role. To date studies have focused on PfAAT1 mutations found in African and Southeast Asian P. falciparum lineages, but chloroquine resistance emerged independently in South America, where there may be novel PfAAT1 polymorphisms that are functionally important. We used AlphaFold modelling to reveal high homology between PfAAT1 and the human lysosomal arginine transporter SLC38A9, which allows prediction of membrane orientation and identifies a partially open channel accessible from the cytoplasm. Several PfAAT1 mutations found only in South American isolates sit near the entrance of this pore, most notably V231 where mutation to aspartate is predicted to alter channel conformation and influence transport, while nearby P446A (which is always found in combination with V231D) and I248T are predicted to impact pore flexibility and local structural stability. To functionally validate these insights, we employed CRISPR/Cas9 gene editing across parasite strains with diverse geographic origins. Reverting the regional V231D mutation in the South American 7G8 strain significantly reduced CQ resistance, providing the first functional evidence that this residue modulates drug susceptibility. Furthermore, introducing the apparently Colombia-specific I248T mutation significantly enhanced parasite multiplication rates in 7G8, demonstrating complex fitness and sensitivity trade-offs. Our findings reinforce the distinct evolutionary trajectory for South American CQ resistance and highlight the necessity of including additional pfaat1 mutations in global molecular surveillance strategies. Author SummaryAntimalarial drug resistance is a major threat to global public health. Chloroquine was used widely in the 1950s-60s as part of a global malaria eradication campaign, but resistance emerged in multiple places independently and chloroquine resistant parasites directly led to the death of millions of children. Chloroquine resistance is primarily driven by mutations in the transporter PfCRT which are thought to increase export of chloroquine from the digestive vacuole, where chloroquine acts to prevent the ability of the parasite to digest haemoglobin as a source of energy. However, it is becoming increasingly clear that additional vacuolar transporters can also modulate chloroquine resistance. In this study we focused on mutations in the putative amino acid transporter 1 (PfAAT1) which are specific to South America, where chloroquine resistance emerged independently from Southeast Asia. By integrating AlphaFold structural predictions with CRISPR/Cas9 gene editing across geographically diverse parasite backgrounds we provide the first functional evidence that the region-specific V231D mutation significantly diminishes chloroquine resistance. These findings emphasise that chloroquine resistance in South America followed a unique trajectory and that PfAAT1 is involved in complex fitness and sensitivity trade-offs. This work emphasises the benefits of integrating diverse experimental and modelling approaches with global molecular surveillance.

Nitrofurantoin is commonly prescribed as a first-line treatment for urinary tract infections (UTIs) and is effective against most uropathogens; however, resistance among Klebsiella spp. remains high. In this study, we investigated synergistic drug combinations--defined as pairs of drugs whose combined effect exceeds that of each agent alone--to enhance treatment efficacy against drug-resistant Klebsiella pneumoniae. A high-throughput drug screen identified six and two small molecules that exhibited >50% synergistic activity in tested strains in combination with nitrofurantoin and ciprofloxacin, respectively. We further validated the top three candidates using nitrofurantoin-susceptible and resistant clinical isolates. Notably, the combination of nitrofurantoin and dequalinium demonstrated strong synergistic activity, observed in 68% of susceptible Escherichia coli strains and 94% of resistant Klebsiella pneumoniae strains tested. Combined inhibition of the citric acid cycle by nitrofurantoin and F1-ATPase by dequalinium resulted in a significant reduction in ATP levels compared to either treatment alone. Importantly, decreased ATP levels did not increase persister cell formation, and dequalinium alone reduced persister cell populations more effectively than nitrofurantoin. However, nitrofurantoin is processed into a poorly understood reactive intermediate whose specific metabolic targets have not been fully elucidated. Using metabolomic analyses, we identified aconitase and isocitrate dehydrogenase--key enzymes in the citric acid cycle--as altered in response to nitrofurantoin. Together, these findings demonstrate that dual targeting of bacterial metabolism by nitrofurantoin and dequalinium represents a promising therapeutic strategy for treating drug-resistant Klebsiella spp. in UTIs.

IntroductionIn sub-Saharan Africa, and more specifically in Cameroon, antimicrobial resistance (AMR) represents a major public health threat. This is underlined by the increasing appearance of multidrug-resistant bacteria. Extended-spectrum {beta}-lactamase producing Escherichia coli (ESBL-Ec), a critical priority bacterium, is increasingly implicated in life-threatening infections in hospital and community settings in Cameroon. Data on the genetic composition of ciprofloxacin-resistant Escherichia coli are limited in Cameroon. This study aimed to investigate the prevalence, genetic diversity, resistance mechanisms in multidrug-resistant Escherichia coli organisms isolated from clinical samples in two hospitals in Yaounde, Cameroon. MethodA cross-sectional study was conducted from February to June 2025 in two healthcare facilities in Yaounde, Cameroon. All clinical samples from in- and out-patients were analysed. After culturing, identification was performed using API20E as per the manufacturers instructions and ESBL production was screened in CHROMagarTM ESBL. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disc diffusion method. Polymerase chain reaction (PCR) was used to detect ESBL and plasmid mediated quinolone resistance (PMQR)genes, as well as mutations in quinolone resistance-determining region (QRDR) (gyrA/parC) Horizontal. plasmid transfer was also investigated. Finally, phylogroup analysis was assessed. ResultThe prevalence of MDR E. coli was 50.7% (n=33/65), all of which (100%) were ESBL producers and 91% were ciprofloxacin-resistant. Highest resistance rates were observed for cefotaxime (100%), ceftriaxone (100%), and ciprofloxacin (91%). The most frequent ESBL genes were blaTEM (36.3%; n=12/33). Among PMQR genes, qnrB was detected in 16.6% (n=5/30) of isolates. Only the ESBL genes were carried by plasmids; the most prevalent plasmid-borne gene was blaTEM (40%), followed by blaCTX-M (26.7%). Mutations within the topoisomerase QRDR (parC gene) were identified in 36.6% (n=11/30) of ciprofloxacin-resistant strains. Phylogroup analysis revealed a predominance of phylogroup A, followed by group B. ConclusionThis study reveals a high prevalence of multidrug-resistance, ESBL (blaTEM dominant) and fluoroquinolone resistance in E. coli in Yaounde, with plasmid dissemination of ESBL genes and chromosomal stabilization of PMQR determinants. The predominance of commensal phylogroups in clinical samples underlines the role of the community reservoir. It is urgent to reinforce " real-time One Health" genomic surveillance in Cameroon.

The human opportunistic pathogen Pseudomonas aeruginosa produces copious quantities of the secondary metabolite pyocyanin (PYO). PYO has recently been shown to interact with the aryl hydrocarbon receptor (AhR), a transcription factor controlling expression of a number of genes, including the cytochrome P450 CYP1A family involved in fluoroquinolone antibiotic clearance. In this study, we investigated whether Pseudomonas aeruginosa could influence the metabolism of ciprofloxacin through human CYP1A2 induction. Primary human hepatocytes were exposed to 1-100 M PYO, with high concentrations resulting in apparent cytotoxicity. Treatment with 5 {micro}M PYO led to a 6.2-fold change in CYP1A2 mRNA and a 3.6-fold increase in oxociprofloxacin metabolite formation. Our results suggest that sub-toxic concentrations of PYO induce CYP1A2 expression and increase ciprofloxacin metabolism, which could lead to sub-efficacious antibiotic concentrations and further drive resistance. This finding directs us to a novel mechanism by which Pseudomonas aeruginosa may escape antimicrobial therapy by hijacking host xenobiotic metabolism pathways. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=110 SRC="FIGDIR/small/740721v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1b350forg.highwire.dtl.DTLVardef@388eb7org.highwire.dtl.DTLVardef@121bc9aorg.highwire.dtl.DTLVardef@1e53060_HPS_FORMAT_FIGEXP M_FIG C_FIG

Antimicrobial resistance (AMR) is a major global public health threat. Wild birds, including seagulls, are increasingly recognised as potential reservoirs and disseminators of resistant bacteria linked to human activity. The objective of the study was to assess the association between human population density and the occurrence of Escherichia coli resistant to critically important antimicrobials in Australian seagulls. Faecal samples were collected from seagull populations in coastal regions across Australia representing differing human population densities. Resistant E. coli isolates were identified and characterised using multilocus sequence typing and plasmid incompatibility group analysis to determine relatedness to human associated lineages. The frequency of resistant E. coli isolation increased with human population density. The predominant sequence types ST10, ST131 and ST354 comprised 24.5% of isolates and belong to globally distributed human associated lineages linked to extraintestinal pathogenic E. coli. Many isolates carried IncF and IncI plasmids, which are key vectors of blaCTX-M extended spectrum beta lactamase genes and plasmid mediated quinolone resistance determinants commonly reported in human clinical strains. IMPORTANCEThese findings support the contention that seagulls primarily acquire resistant bacteria through contact with anthropogenic activities. Once acquired, these bacteria may be disseminated to other seagulls, birds and animals, including being transmitted to humans.

Pharmacodynamic curves describe how changes in drug concentration affect pathogen growth. They are essential for designing treatments that promote pathogen eradication and minimize the evolution of antibiotic resistance. The classical function for modelling pharmacodynamics is a phenomenological, S-shaped curve with stable growth and death rates separated by a single drop. In this study, we characterized the pharmacodynamic curve of the {beta}-lactam antibiotic cefotaxime (CTX) acting against Escherichia coli. We found that the relationship between CTX concentration and net growth rate diverged from classical model predictions, instead yielding a two-step curve defined by distinct phases of growth, population maintenance, and killing. We hypothesized that the intermediate phase arose from antibiotic tolerance conferred by bacterial filaments. Microscopic assessment of treated cells indeed showed a difference in degree of filamentation with concentration. We further sought to explain this with a semi-mechanistic pharmacodynamic function, modelling the binding of CTX to its cellular targets, penicillin binding proteins (PBP) 1 and 3. By incorporating the preferential concentration-dependent binding of CTX to PBP3 and then PBP1, yielding filaments or lysed cells respectively, we replicated the two-step curve in silico. We also assessed the pharmacodynamics of CTX against mutants conferring resistance; these displayed further altered curves, in line with their fitness costs. Altogether, our results show that CTX has a two-step pharmacodynamic curve against E. coli arising from multiple targets separated in their affinity for the antibiotic. We present a model offering a mechanistically grounded framework for capturing such dynamics. These pharmacodynamic curves deserve careful consideration when defining optimal dosing.

Background: Antimicrobial resistance (AMR) disproportionately affects low- and middle-income countries (LMICs) such as Pakistan, where obstetric and gynaecological (OBGYN) patients carry high antibiotic exposure. Specialty-specific drug utilization data with concurrent stewardship audit remain scarce. This study evaluated antibiotic prescribing patterns, consumption metrics, and antimicrobial stewardship program (AMS) compliance in OBGYN inpatients at a public sector tertiary care hospital. Methods: A prospective cross-sectional study was conducted in OBGYN wards of Dow University Hospital, Karachi, from 1 September to 31 October 2025. Women receiving [&ge;]1 systemic antibiotic were included. Daily AMS rounds were conducted by an Infectious Diseases physician and pharmacist. Antibiotic consumption was measured as Defined Daily Doses (DDD) and Days of Therapy (DOT) per 1,000 patient-days (total = 821). Antibiotics were classified by WHO AWaRe (2023) framework. Results: Of 812 total admissions, 278 patients (34.2%) received [&ge;]1 antibiotic and were enrolled (205 obstetric, 73 gynaecological), generating 636 prescriptions (mean 2.29/patient). Surgical prophylaxis was the predominant documented indication (213, 33.5%); 65.1% carried no documented indication. By AWaRe classification, 53.6% were Access-group and 46.1% Watch-group. Ceftriaxone (38.4%) and metronidazole (36.8%) together represented 75.2% of prescriptions. Combined DDD/1,000 patient-days was 1,758.6 and DOT/1,000 patient-days was 1,852.7. AMS compliance was 0%. Conclusions: This study documents high antibiotic prescribing burden, near-universal documentation failure, and zero AMS compliance in OBGYN inpatients at a Pakistani public sector hospital. The predominance of Watch-group antibiotics and undocumented surgical prophylaxis highlights structural stewardship gaps. Findings support urgent need for institutional OBGYN antibiotic guidelines and structured pharmacist-led AMS programs.