Combined exposure to thermal and antibiotic stresses is ubiquitous. How cells adapt to these concurrent cues at the single-gene level remains unresolved. Here, we show that in Escherichia coli, both cold and heat shock responses dominate antibiotic stress responses across multiple levels of physiology. Using RNA-seq, we demonstrate that combined exposure produces transcriptomes that are largely indistinguishable from those induced by thermal shifts alone. Flow-cytometry measurements across intermediate temperatures and increasing ofloxacin concentrations further show that temperature-like responses are more prevalent outside the 30°C-39°C range. This dominance arises from temperature-driven changes in ATP levels and nucleoid organization, which in turn modulate DNA gyrase and RNA polymerase engagement with DNA. Combined exposure also induces SoxS- and MarA-mediated efflux systems more strongly than individual stresses. Machine learning analysis identifies CRP and σ38 as key gene expression regulators under combined stress. Molecular dynamics simulations show that drug-target conformational plasticity is altered under combined conditions. Cross-species simulations across six bacterial species with distinct pathogenic profiles reveal that thermal modulation of antibiotic response is partially conserved across evolutionarily distant species. Together, the results identify temperature as a critical determinant of antibiotic response, suggesting that thermal shifts may facilitate bacterial adaptation to antibiotics.

Trace residues of fluoroquinolones (FQs) in food matrices are difficult to determine directly because of their low concentrations and the complexity of protein- and sugar-rich samples. In this work, cotton fibers functionalized with a TAPB-DHTP covalent organic framework (TAPB-DHTP COF@cotton) were prepared as a packed adsorbent for in-syringe solid-phase extraction (IS-SPE) of FQs. (3-aminopropyl)triethoxysilane (APTES) was first grafted onto cotton fibers through condensation between the ethoxy groups of APTES and the surface hydroxyl groups of cotton, introducing amino groups onto the fiber surface. TAPB-DHTP COF was then grown in situ on the APTES-modified fibers through Schiff-base condensation between TAPB and DHTP. This design combines the easy packing and flexibility of cotton fibers with the rough surface and adsorption sites provided by the COF coating. The obtained material was characterized using several instrumental techniques. After coupling with ultra-performance liquid chromatography, the method was used for the determination of enoxacin, levofloxacin, and ciprofloxacin. Under the optimized extraction conditions, the method showed good linearity over 0.5-100 ng/mL, with correlation coefficients of 0.9991-0.9996. The limits of detection were 0.02-0.08 ng/mL, and the intra-batch and inter-batch relative standard deviations (RSDs) were below 4.66% and 4.85%, respectively. The extraction efficiencies and enrichment factors were 76.9-83.8% and 51.31-55.84. The method was applied to honey, egg white, and fish samples, giving recoveries of 81.05-97.23% with RSDs below 9.77%. The results demonstrate that TAPB-DHTP COF@cotton provides a practical fibrous adsorbent for trace fluoroquinolone determination in complex food matrices.

Aquatic antibiotic pollution represented by ofloxacin (OFX) causes serious ecological hazards and endangers public health due to the high persistence and bioaccumulation of antibiotic residues. Conventional water treatment techniques are insufficient for OFX elimination, limited by low removal efficiency, high energy consumption, and poor operational stability. Herein, a novel MoS2/CuS heterojunction composite was fabricated via a hydrothermal method and applied to an ultrasound-driven piezocatalysis-coupled peroxymonosulfate (PMS) advanced oxidation system for OFX wastewater remediation. The introduction of CuS effectively remedies the inherent shortcomings of pristine MoS2, including insufficient active sites and rapid photogenerated carrier recombination. The constructed heterojunction induces a strong interfacial built-in electric field, which significantly accelerates the migration of piezoelectric charges. The synergistic photo-piezoelectric effect further promotes continuous PMS activation and facilitates the massive generation of reactive oxygen species (ROS). The influences of key operating parameters and common water inorganic anions on OFX degradation performance were systematically investigated. Radical trapping experiments confirmed the synergistic mechanism between piezocatalysis and PMS activation during the catalytic reaction. The optimized MoS2/CuS heterojunction exhibits remarkable OFX degradation efficiency and excellent cyclic stability. This work provides a feasible strategy for the rational design and fabrication of high-efficiency piezocatalysts and offers a promising technical route for the remediation of refractory antibiotic wastewater via piezocatalysis-coupled PMS advanced oxidation.

Polymicrobial fracture-related infections (FRIs) underpinned by bacterial biofilms pose therapeutic challenges distinct from those of monomicrobial infection, as interspecies dynamics may alter both antimicrobial response and host tissue injury. This study evaluated systemic and local antibiotic strategies in an ovine FRI model co-inoculated with a polymicrobial Staphylococcus aureus and Pseudomonas aeruginosa biofilm. At 21 days, untreated positive controls retained substantial total bioburden yet paradoxically preserved bone volume and exhibited low osteoclast activity, contrasting with the marked osteolysis previously observed in monomicrobial infection controls using this model. Although untreated controls remained S. aureus predominant, their S. aureus:P. aeruginosa ratio declined from approximately 200:1 at inoculation to 15:1, indicating a progressive ecological shift over time. A systemic levofloxacin plus rifampin regimen more effectively reduced S. aureus than P. aeruginosa. Likewise, all antibiotic-treated groups shifted from an S. aureus-dominant inoculum toward P. aeruginosa predominance, consistent with unequal antimicrobial suppression altering polymicrobial pathogen balance. Single-dose local antibiotic powder and calcium sulfate bead strategies provided inconsistent microbiological benefit and failed to preserve bone. Reloadable local antibiotic delivery reduced S. aureus tissue bioburden and preserved the greatest bone volume among antibiotic-treated groups but the antibiotics selected for delivery did not prevent persistent P. aeruginosa colonization or the shift in pathogen balance. These findings indicate that microbiological burden reduction, residual pathogen composition, and bone preservation may result in distinct treatment outcomes in polymicrobial FRI. This study highlights the need for appropriate antibiotic selection with ecologically informed, dual targeted local and systemic strategies to manage polymicrobial, biofilm-complicated FRIs.

Microbacterium species, a genus of aerobic Gram-positive coryneform rods, are rare causes of human infection with only a few sporadic cases reported worldwide. MALDI-TOF MS, 16S rRNA gene sequencing, and even whole-genome sequencing are used to identify the species in a clinical laboratory setting. The study reports a case of Microbacterium paraoxydans bloodstream infection and reviews the literature, including eight additional reported cases of M. paraoxydans infection. We summarize the characteristics of each case, highlight the identification methods, and identify recurrent characteristics for M. paraoxydans infection. A 65-year-old man presented with stable condition of esophageal cancer and long-term prednisolone therapy for psoriasis. A central venous catheter (CVC) was placed for years for treatment of esophageal cancer. The patient presented with fever and chills, and Microbacterium paraoxydans was identified by MALDI-TOF MS from blood culture samples via chemo port. After the chemo port was removed, vancomycin followed by levofloxacin was administered for the entire 14-day treatment course according to susceptibility tests. This study highlights that a long-term central venous catheter and immunocompromised status were common characteristics among the reported cases of Microbacterium spp. infection. The susceptibility of Microbacterium spp. is not fully characterized and varies across reports; gentamicin and trimethoprim/sulfamethoxazole appeared more consistently active, whereas susceptibility to vancomycin was variable. We report this case to highlight the increasing number of infections caused by Microbacterium spp. in the current era and to provide information on the treatment of M. paraoxydans.

Antimicrobial resistance (AMR) is one of the main public health problems that also affects environmental ecosystems. Sinkholes are understudied groundwater ecosystems that may act as reservoirs and dissemination routes for AMR. This study evaluated the composition, distribution, and antimicrobial resistance profiles of culturable ampicillin-resistant Gram-negative bacteria recovered from sinkholes of the Yucatán Peninsula, Mexico. Water samples were collected from 27 sinkholes during dry and rainy seasons. The isolates were identified by MALDI-TOF MS and evaluated by broth microdilution. Representative E. coli isolates were analyzed by whole-genome sequencing. A total of 193 ampicillin-resistant Gram-negative isolates were recovered. The most frequently recovered genera were Enterobacter, Klebsiella, and Escherichia, which were detected across all hydrogeological zones. Resistance to cefotaxime, levofloxacin, gentamicin, and tetracycline was detected among several bacterial genera. Opportunistic bacterial species associated with healthcare-associated infections, including K. pneumoniae, A. baumannii, and P. aeruginosa, were also identified. Genomic analysis of 19 E. coli isolates revealed resistance determinants associated with multiple antimicrobial classes and a diverse population structure dominated by phylogroup B1. These findings show that the Ring of Cenotes aquifer harbors diverse populations of culturable ampicillin-resistant Gram-negative bacteria and emphasize the importance of groundwater ecosystems as environmental reservoirs of AMR.

One of the most important adverse consequences of antibiotic use is the development of bacterial resistance. This study investigated the prevalence and antimicrobial resistance patterns of multidrug-resistant (MDR) Escherichia coli in a tertiary care hospital during 2013-2015 and 2024, together with trends in antibiotic consumption and the molecular characteristics of resistance genes in 2024 isolates. Identification and susceptibility testing were performed using the Vitek® 2 system, antibiotic consumption was assessed according to WHO ATC/DDD methodology, and resistance genes were detected by PCR. No significant differences were observed in the total number of isolates, patients, or the proportion of E. coli isolates during 2013-2015. However, the isolation rate of MDR E. coli significantly declined from 36.9% to 30%. Total antibiotic consumption remained stable, with no correlation between consumption and MDR isolation rates, although ampicillin resistance increased significantly. In 2024, MDR E. coli accounted for 24.3% of isolates despite lower antibiotic consumption. Compared with 2015, resistance significantly increased to amoxicillin-clavulanic acid, cefotaxime, cefepime, ceftazidime, ciprofloxacin, and levofloxacin, while ceftriaxone resistance decreased. The blaCTX-M gene was detected in 53% of bloodstream isolates, indicating widespread dissemination of ESBL-producing E. coli and highlighting the need for continuous surveillance and antimicrobial stewardship.

Detection systems for antibiotics of the (fluoro)quinolone (FQ) group are in high demand among food safety regulators and agricultural inspectors. At the same time, the application of developed test systems may be complicated by the significant structural diversity of related FQs (including enantiomeric forms), which necessitates the careful evaluation of their selectivity. In this study, five polyclonal antibodies were generated by immunizing rabbits with the S- and R-enantiomers of ofloxacin (OFL) and its racemic mixture used as haptens. The cross-reactivity (CR) of the obtained antibodies toward 26 FQs, structural analogs of OFL, was evaluated using an indirect competitive enzyme-linked immunosorbent assay. Structure-activity relationship models were developed to analyze the obtained CR data. Three machine learning (ML) methods were applied: random forest classifier (RFC), logistic regression (LR), and a support vector classifier (SVC). The SVC model demonstrated the highest predictive performance in terms of the Log Loss metric. In contrast, the LR models showed the best overall balance across six statistical metrics, including precision, recall, and F1 score. Three-dimensional topological descriptors enabled discrimination between the S- and R-isomers of OFL, whereas constitutional and two-dimensional descriptors were less effective. The obtained results contribute to the development of FQ immunoassay systems and their computational analysis using ML approaches.

Background/Objectives: Antibiotic-associated encephalopathy (AAE) is a serious adverse reaction whose presentation is thought to differ between antibiotics; whether these agent-specific profiles are consistently reflected in spontaneous reports across independent data sources is unknown. Methods: Using the Japanese Adverse Drug Event Report (JADER) database and the US FDA Adverse Event Reporting System (FAERS, 2004-2026), we studied metronidazole, cefepime, ceftriaxone, ceftazidime, and levofloxacin. Encephalopathy was defined by the MedDRA terms Encephalopathy or Toxic encephalopathy, and reporting odds ratios (RORs) were calculated in each database; co-reported neurological manifestations, time to onset, patient age, and renal-associated terms were then characterized. Results: All five antibiotics showed significant reporting disproportionality in both databases, with metronidazole and cefepime showing the largest RORs in each (JADER: metronidazole ROR 186.70, cefepime 55.33; FAERS: cefepime 69.29, metronidazole 29.11), consistent with established adverse reactions. In FAERS, metronidazole showed a cerebellar profile (ataxia OR 12.48, dysarthria OR 11.11) and cefepime a renal-associated altered-mental-status/myoclonus profile (depressed level of consciousness OR 12.88, myoclonus OR 11.76; renal-associated terms OR 2.88; median age 71 years); event-specific JADER dates indicated a delayed course for metronidazole (median 28 days). The symptom-level contrast was FAERS-based and was not reproduced when the same analysis was repeated in JADER, a discrepancy that may reflect differences in coding practice. Conclusions: AAE is reported not as a single uniform event but with profiles specific to the causative antibiotic. Awareness of these agent-specific patterns may support earlier recognition, although confirmation in analytic studies with exposure denominators is warranted.

Background/Objectives:Streptococcus parasuis is an under-recognized member of the Streptococcus suis complex that can be misidentified by routine diagnostic methods. Human clinical isolates remain sparsely characterized. This study investigated the antimicrobial phenotype, genomic features, population position, and larval pathogenicity of a human pleural-fluid isolate. Methods: Strain HUGSPH was recovered during the care of a patient with pneumonia. Hybrid whole-genome sequencing (WGS), average nucleotide identity analysis, multilocus sequence typing (MLST), core-genome phylogenetics, comparative pan-genomics, and an exploratory host-origin association analysis were performed. Minimum inhibitory concentrations (MICs) were determined using the VITEK 2 Compact system with an AST-ST03 card, and virulence was evaluated in Galleria mellonella larvae. Results: Genome-based analysis confirmed HUGSPH as S. parasuis and assigned it to the novel sequence type (ST) 181. The isolate clustered within a predominantly human-origin phylogenetic clade. Because species-specific clinical breakpoints are unavailable, the MICs were interpreted provisionally using explicitly stated surrogate Clinical and Laboratory Standards Institute (CLSI) criteria; erythromycin, clindamycin, and levofloxacin were categorized as resistant under those criteria. Several predicted antimicrobial-resistance-associated genes were detected. In a restricted comparison of seven human-origin and nine swine-origin genomes, eight genes were present in all included human-origin isolates and absent from all included swine-origin isolates, whereas metQ, metP, and metN showed the reciprocal pattern. HUGSPH caused dose-dependent larval mortality, reaching 100% at 107 colony-forming units (CFU) by 96 h. Conclusions: HUGSPH expands the genomic record of human clinical S. parasuis and highlights the diagnostic and surveillance relevance of antimicrobial resistance in this species. The susceptibility categories, source-associated genes, and larval phenotype require validation by reference susceptibility testing, broader phylogenetically balanced collections, comparative strains, and mammalian models.

The Burkholderia cepacia complex (BCC) is comprised of 24 species of gram-negative bacteria that cause opportunistic infections. While antimicrobial susceptibility testing (AST) has historically been used to guide treatment for BCC infections, recent work highlighting problems with AST for these organisms led the Clinical and Laboratory Standards Institute (CLSI) to remove disk diffusion and minimal inhibitory concentration (MIC) breakpoints for BCC from its M100 standards document. Epidemiological cut-off values (ECVs) may be helpful to clinicians in the absence of breakpoints, as they may be used to determine whether an isolate has a wild-type or non-wild-type phenotype. Here, we present an analysis of BCC ECVs for ceftazidime (CAZ), levofloxacin (LVX), meropenem (MEM), minocycline (MIN), and trimethoprim-sulfamethoxazole (TMP-SMX). ECVs were calculated using MIC data from 3 previous studies and 3 independent laboratories for 1,896 BCC isolates. ECVs were 16 μg/mL for CAZ, 8 μg/mL for LVX, 16 μg/mL for MEM, and 8 μg/mL for MIN. The ECV for TMP-SMX varied depending on the analysis from 2, 8, and 16 μg/mL and therefore could not be reliably established. Challenges with establishing ECVs for BCC include limitations with the pooled MIC data set, broad MIC distributions, and high ECVs that are above the obsolete susceptible MIC breakpoints. These challenges limit the clinical utility of ECVs for these organisms and support the removal of ECVs from the CLSI M100 standards document.