Therapeutic option for treating methicillin-resistant Staphylococcus aureus (MRSA) infection is urgently required since its resistance to a broad spectrum of currently available antibiotics. Here, we report that isoniazid is able to potentiate the killing efficacy of tigecycline to MRSA. The combination of isoniazid and tigecycline reduces the minimal inhibitory concentration of clinic MRSA strains to tigecycline. The killing activity of tigecycline is further confirmed by killing experiments and murine infection model. We further demonstrate the mechanism that isoniazid increases intracellular accumulation of tigecycline by promoting the influx but limiting the efflux of tigecycline through proton motive force. We also show that isoniazid and tigecycline synergize to increase the abundance of isoniazid-NAD adduct, which in turn damage cell membrane, possibly contributing to the disruption of PMF. Whereas phosphatidylethanolamine and cardiolipin are able to abrogate the synergistic effect of isoniazid plus tigecycline. Thus our study provides a new perspective that antibiotics, e.g. isoniazid, once recognized only to target Mycobacterium tuberculosis, can be repurposed as antibiotic adjuvant to tigecycline, expanding our choice of antibiotic-antibiotic combinations in treating bacterial infectious diseases.

Novel benzothiazole aryl ureas were designed and synthesized as anti-MRSA agents targeting peptidoglycan (PG) hydrolases (autolysins). Structural simplification of prior benzothiazole-urea hybrids yielded compounds 4a, 7a and 11a bearing p-CF3 on phenyl ring demonstrating narrow-spectrum activity against Gram-positive bacteria including clinical methicillin-resistant S. aureus (MRSA). The primary autolysin in S. aureus, AtlA, mediates peptidoglycan hydrolase activity critical for bacterial growth, division, and cell wall remodeling. Mechanistic studies revealed that 4a down-regulated autolysin-related genes RNAIII and walR, disrupting peptidoglycan homeostasis. Knockout of atlA (a key autolysin gene) impaired 4a's efficacy, confirming autolysins as critical targets. Docking indicated that 4a binds to AtlA via hydrogen bonds, Pi-Pi, and hydrophobic interactions. In vivo, 4a significantly reduced bacterial load in a murine abdominal infection model, outperforming vancomycin at 10 mg/kg with lower cytotoxicity. Additionally, 4a disrupted MRSA biofilms, suppressed hemolytic toxin production, and alleviated inflammation in infected mice. These findings underscore AtlA as a promising therapeutic target and highlight benzothiazole phenyl urea as a scaffold for developing innovative anti-staphylococcal agents.

Staphylococcus aureus, a gram-positive bacterium, causes infective endocarditis, osteoarticular, skin, and respiratory infections. The emergence of multidrug-resistant strains, particularly Methicillin-resistant Staphylococcus aureus (MRSA), has caused a 21-35 % rise in bloodstream infections, complicating treatment strategies. Filamentous temperature-sensitive protein Z (FtsZ), a critical protein involved in bacterial cell division, forms a Z-ring at the division site, making it a key target for novel antibacterial therapies. In this study, 1165 phytochemicals were screened, and three lead molecules namely, Aromadendrin, Leucopelargonidin, and 7-Deacetoxy-7-oxogedunin were identified based on their favorable physicochemical properties, drug-likeness, and estimated binding affinities (- 11.73 kcal/mol, - 10.77 kcal/mol, and - 10.38 kcal/mol, respectively) against FtsZ. 100 ns Molecular dynamics simulations conducted in triplicates confirmed the stability of the FtsZ-ligand complexes.Binding free energy calculations revealed that IMPHY003535 (Leucopelargonidin) exhibited the most favorable binding free energy (-27.25 kcal/mol), followed by 7-Deacetoxy-7-oxogedunin (-15.31 kcal/mol) and Aromadendrin (-13.38 kcal/mol). Leucopelargonidin emerged as the most promising inhibitor, highlighting its potential as a lead compound for developing antibacterial agents targeting FtsZ. These findings demonstrate the significant role of phytochemicals in combating antibiotic resistance and the importance of further optimization, including in vivo studies, to assess their therapeutic potential, which could provide new treatment avenues to overcome bacterial resistance mechanisms.

The globally disseminated Staphylococcus aureus ST5 clone poses a major public health threat due to its multidrug resistance and virulence. Here, we identified an agr-dysfunctional (agrA-I238K) ST5 MRSA clone that has spread across East and Southeast Asia, with recent increases in China since its emergence in the 1970s. Comparative genomic analyses identified distinct single-nucleotide polymorphisms and mobile genetic elements linked to enhanced resistance and virulence. This clone exhibits resistance to seven antimicrobial classes, including third-generation tetracyclines and fusidic acid, and shares phenotypic and genetic similarities with the vancomycin-intermediate S. aureus Mu50 strain, including reduced susceptibility to vancomycin, teicoplanin, and daptomycin. The agrA-I238K mutation attenuates hemolytic activity, increases biofilm formation, and reduces daptomycin susceptibility, suggesting a key role in the clone's success. Our results demonstrate the important role of agrA-I238K mutation in the widespread distribution of agr-dysfunctional MRSA and highlight the importance of genomic surveillance in tracking the spread of agr-dysfunctional ST5 MRSA.

The management of infected chronic wounds is one of the urgent challenges. Herein, a hesperidin (Hes)-loaded self-assembled supramolecular hydrogel based on quaternized chitosan (CQHP) has been developed as an efficient photothermal antibacterial dressing for MRSA-infected wound healing. Specifically, CQHP hydrogels fabricated through the dynamic noncovalent interactions among CM-β-CD grafted QCS, Hes, proline and Fe3+, exhibited injectable and self-healing behaviors, along with adhesion, antioxidant, hemostatic and protein adsorption performance, satisfying the essential feature as chronic wound dressing. Of note, superior photothermal effect generated from the Hes-Fe3+ has been demonstrated, which endowed the CQHP hydrogels effectively and rapidly eliminate the E. coli, S. aureus and MRSA through photothermal therapy, thereby avoiding the use of antibiotics or photothermal conversion nanomaterials in hydrogels and substantially reducing the biological toxicity. Furtherly, sustained antibacterial performance in the absence of NIR can be achieved through the inherent antibacterial activities of Hes. Importantly, the developed CQHP hydrogels significantly promoted the closure of acute full-thickness scratch wounds, and exhibited remarkable better therapeutic effect on MRSA-infected wound than commercial 3M transparent film, by efficiency and sustained antibacterial activity, reducing inflammation, enhancing angiogenesis and collagen deposition, highlighting its promising application in the MRSA-infected wound healing with high efficiency, quality and security.

Superficial surgical site infection (SSI) is a significant risk factor for the development of periprosthetic joint infection (PJI), particularly in diabetic patients. A high-glucose microenvironment is observed to compromise phagocytosis by inducing cellular senescence, which leads to impaired antibacterial immune function. Exosomes derived from umbilical cord stem cells (H-Exos) can reverse the immunosuppressive microenvironment by rejuvenating senescent cells, thereby terminating excessive, persistent, and ineffective inflammatory responses. Thus, a novel exosome-based immunotherapeutic antibacterial strategy to reverse fate is proposed. Vancomycin & lysostaphin-loaded exosomes are incorporated in a customizable microneedle patch (ExoV-ExoL@MN) for controlled release, enabling tailored treatments for diverse clinical scenarios. While rejuvenating macrophage senescent phenotype, the antibiotics encapsulated within exosomes can be responsively released by the hemolysin secreted by bacteria, triggering rapid bacterial killing. Post-infection clearance, they induce a shift from M1 to M2 macrophage polarization, thereby enhancing anti-inflammatory and reparative responses. Furthermore, the components can be mixed on demand and at any time, allowing for real-time customization and fabrication directly at the clinic (fabrication@clinic). This strategy reverses the immunosuppressive microenvironment by rejuvenating senescent macrophages and effectively combats bacterial invasion into deep tissues through bacteria-responsive antibiotic release, providing a promising approach for preventing and treating SSI-induced PJI.

The human defensins are a group of cationic antimicrobial peptides that range in size from 2 to 5 kDa and share a common structural motif of six disulphide-linked cysteines. Several naturally occurring human α- and β-defensins have been identified over the past two decades. They have a wide variety of antimicrobial effects, and their potential to avoid the development of resistance to antimicrobial treatment makes them attractive as therapeutic agents. Human defensins have recently been the focus of medical and molecular biology studies due to their promising application in medicine and the pharmaceutical industry. This work aims to provide a comprehensive summary of the current developments of human defensins, including their identification, categorization, molecular features, expression, modes of action, and potential application in medical settings. Current obstacles and future opportunities for using human defensins are also covered. Furthermore, we shed light on the potential of this class as an antiviral agent, particularly against SARS CoV-2, by providing an in silico-based investigation of their plausible mechanisms of action.

Bacterial resistance is a major challenge in the treatment of Staphylococcus aureus infections, with efflux mechanisms highlighted as reducing the efficacy of antibiotics. In this study, we investigated the potential of naringenin, a natural flavonoid, as an antibacterial agent and efflux pump inhibitor in S. aureus strains 1199 and 1199B. The studies used minimum inhibitory concentration (MIC) assays, ethidium bromide (EtBr) fluorescence emission enhancement assays, cell membrane permeability assays, and in silico molecular docking and ADME prediction assays. Naringenin showed no relevant antibacterial activity (MIC ≥1024 μg/mL). However, it potentiated the effect of norfloxacin and EtBr, reducing their MICs and increasing the fluorescence emission of EtBr, suggesting a possible inhibition of the NorA efflux pump. Bacterial membrane permeability was not significantly affected. Molecular docking assays indicated that naringenin interacts with the chlorpromazine binding site and has more favorable affinity energy than the chlorpromazine-NorA complex. ADME prediction showed favorable physicochemical properties, good oral absorption, metabolic stability and central nervous system safety. Therefore, naringenin demonstrates the potential to reverse the efficacy of norfloxacin in S. aureus by associating with efflux inhibition through effective interactions with the NorA protein, suggesting its therapeutic potential against bacterial resistance.

MRSA is one of the main pathogens that cause nosocomial pneumonia. Based on longitudinal in vitro and in vivo data, a pharmacokinetic-pharmacodynamic (PKPD) model was built to quantify the effect of two control antibiotics (LZD and VAN) for Gram-positive bacteria in a standardized mouse pneumonia model.

The PKPD model was developed for data generated on the MRSA strain 160 079 in static in vitro time-kill experiments and thereafter adjusted to fit data from lungs of neutropenic mice administered with single or multiple doses of LZD (0.5-40 mg/kg) or VAN (1-40 mg/kg). Simulations with human PK were run to predict antibacterial response in patients.

Bacterial regrowth observed in vitro when exposed to VAN concentrations was described by an adaptive resistance model. The selected MRSA isolate showed good virulence in the mouse pneumonia model. Bacterial load in lungs decreased up to 2-log with respect to control mice after LZD and VAN treatment. A 70%-75% lower killing rate was estimated for the in vivo data when compared with in vitro. Simulations displayed bacterial stasis at 24 h for patients infected with bacteria with MICs below the clinical breakpoint for both drugs after administering standard-of-care dosing regimens.

A translational workflow allowed us to build a PKPD model with both in vitro and in vivo data that characterized bacterial dynamics following LZD and VAN exposure, showing that this approach can inform the development of antibiotics. We also showcased the first successful use of the standardized mouse pneumonia model for Gram-positive bacteria.

Methicillin-resistant Staphylococcus aureus (MRSA) has spread across diverse global environments, and MRSA-related infection is a major threat to public health. Implant-associated infection (IAI) caused by MRSA remains a tough global clinical problem. Conventional antibiotic therapy has limited efficacy in treating MRSA-related IAI, and antibiotic abuse has resulted in the emergence of multidrug-resistant bacteria. Hence, there is a necessity to explore more effective approaches to deal with MRSA-related IAI. Herein, inspired by neutrophil extracellular traps (NETs) released by neutrophils to kill microorganisms, this study proposes a novel biomimetic nano-NET strategy using an epsilon-poly-l-lysine-coated CuO2 nanoplatform, denoted as PCPNAs. The function-adaptive nanoplatform exhibited excellent Fenton-like performance, including robust ROS generation and GSH scavenging ability. PCPNAs showed >90% cell viability in mammalian cells and reduced bacterial burden by 7.65 log10 CFU in vitro. Moreover, the positively charged PCPNAs could easily bind to negatively charged MRSA cells through charge-coupling and simultaneously exerted a trapping effect on MRSA cells. Notably, PCPNAs self-assembled into web-like structures to physically trap and kill biofilm bacteria, achieving 99.58% biofilm eradication. Furthermore, PCPNAs showed satisfactory biocompatibility in vivo and displayed ideal anti-bacterial and anti-inflammatory effects in a mouse model with implant-associated infection. With further development and optimization, the biomimetic nano-NET strategy based on PCPNAs provides a new therapeutic option for the treatment of MRSA-related implant-associated infection.

Antimicrobial resistance (AMR) is one of the leading health crises worldwide that demands new antimicrobials to enter the clinical pipeline. Marine sponges are a rich source of promising bioactive compounds. Due to their sessile nature and filter-feeding lifestyle, sponges are prone to attack by competitors, predators, and pathogens. To combat these threats, they produce a diverse array of bioactive compounds. Notably, the microbial communities residing within the sponges make many of these beneficial compounds. Twenty-one bacterial isolates from various marine sponges from the Indian coast were selected for this study. The bacterial isolates were fermented to obtain crude extracts, which were then screened against critical bacterial pathogens. Based on the MIC (minimum inhibitory concentration) results, two isolates, Bacillus velezensis NIO_002 and Stutzerimonas stutzeri NIO_003 showing good activity, were characterized by morphological, biochemical, and molecular methods. Genome mining predicted multiple antibiotic biosynthetic gene clusters, most of which showed a high degree of similarity to known gene clusters, and some with low or no similarity which may be indicative of novel gene clusters. LC-MS (liquid chromatography-mass spectrometry) data revealed the putative presence of certain antibacterial compounds previously reported in the literature. To our knowledge, this is the first study to report the antimicrobial activity of marine sponge-associated Bacillus velezensis and Stutzerimonas stutzeri strains characterized by whole genome sequencing, thereby indicating the novelty of our strains. This study emphasizes the potential of our bacterial isolates for further development as a source of promising antibiotics to address the escalating challenge of drug-resistant pathogens.

Bacteria with similar genomes can exhibit different phenotypes due to alternative gene expression patterns. In this study, we analysed four antibiotic-resistant Staphylococcus aureus hospital isolates using transcriptomics, PacBio genome sequencing, and methylomics analyses. Transcriptomic data were obtained from cultures exposed to gentamicin, the iodine-alanine complex CC-196, and their combination. We observed strain-specific expression patterns of core and accessory genes that remained stable under antimicrobial stress - a phenomenon we term the Clonal Gene Expression Stability (CGES) that is the main discovery of the paper. An involvement of epigenetic mechanisms in stabilization of the CGES was hypothesized and statistically verified. Canonical methylation patterns controlled by type I restriction-modification systems accounted for ~ 10% of epigenetically modified adenine residues, whereas multiple non-canonically modified adenines were distributed sporadically due to imperfect DNA targeting by methyltransferases. Protein-coding sequences were characterized by a significantly lower frequency of modified nucleotides. Epigenetic modifications near transcription start codons showed a statistically significant negative association with gene expression levels. While the role of epigenetic modifications in gene regulation remains debatable, variations in non-canonical modification patterns may serve as markers of CGES.

S-scheme heterojunctions have garnered significant attention in the field of photocatalytic antimicrobials due to their superior charge separation efficiency and higher redox capacity. In this study, an innovative linear conjugated polymer (PCO) was combined with fragmented carbon nitride (TP-PCN) to create PCO/TP-PCN organic-organic S-scheme heterojunctions, which markedly enhanced the photocatalytic antimicrobial performance. The composite (PCO-7/TP-PCN) demonstrated the ability to combat bacterial infections under visible light irradiation, effectively eradicating approximately 2.16 × 107 cfu/ml MRSA within 6 min. This exceptional photocatalytic performance can be attributed to the successful formation of an S-scheme heterojunction between PCO and TP-PCN, as well as the interaction of surface functional groups of PCO-7/TP-PCN with bacteria. Results from UV-Vis-NIR DRS and in situ-XPS experiments indicated a significant enhancement in carrier transport rate through the establishment of a built-in electric field and energy band bending at the interface. In vitro and in vivo experiments further demonstrated that PCO-7/TP-PCN not only exhibited potent antimicrobial activity under visible light irradiation but also promoted angiogenesis to inhibit inflammatory responses. Therefore, it can be concluded that this organic-organic S-scheme heterojunction photocatalyst holds great potential for development as a promising new generation of efficient antimicrobial materials, which could aid in preventing bacterial infection of wounds and ensuring effective wound healing.

Diabetic wounds (DWs) are characterized by high blood glucose levels, and one of the primary strategies for regulating blood glucose is the use of glucose oxidase (GOx). This enzyme catalyzes the oxidation of glucose to produce D-gluconic acid, consuming oxygen and generating hydrogen peroxide (H₂O₂) in the process. In DWs, this reaction not only effectively reduces glucose concentrations at the wound site but also provides an antibacterial effect through the release of H₂O₂. Based on this principle, combining glucose oxidase with other therapeutic approaches to develop multimodal wound treatment strategies has garnered significant research attention. Additionally, the abundance of binding sites on the GOx molecular surface enables the construction of multifunctional GOx-based nanocomposites. This review uniquely integrates emerging nanomaterial designs with cascade therapeutic strategies, offering insights into overcoming challenges in diabetic wound healing. Recently, multifunctional nanocomposites have gained attention for integrating multiple therapeutic modalities, relying on cascade mechanisms of multimodal synergistic therapies to tackle complex challenges in DWs treatment. However, there is currently no systematic review that comprehensively elaborates on the construction of these nanocomposites and the specific applications of multimodal treatment strategies in DWs healing. To fill this gap in the field, this review provides a comprehensive overview of these nanomaterials, starting with a systematic exploration of cascade and synergistic therapeutic mechanisms centered on GOx-catalyzed reactions. It highlights applications in photothermal therapy (PTT), photodynamic therapy (PDT), and gas therapy (GT), summarizes the design of nanocarriers, and discusses challenges in DWs healing and future development directions. The findings discussed provide a pathway for the development of clinically viable, cost-effective therapies for chronic wounds.

Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant global health challenge, particularly associated with serious infections such as bacteremia. Lipoglycopeptide antibiotics, including vancomycin, dalbavancin, and daptomycin, are critical in MRSA treatment. In this study, we analyzed two MRSA isolates (XF1 and XF2) from a bacteremia patient treated with vancomycin. Antimicrobial susceptibility testing revealed that XF1 was sensitive to vancomycin, dalbavancin, and daptomycin, whereas XF2 exhibited 8- to 16-fold higher minimum inhibitory concentrations for these antibiotics, alongside a 4- to 8-fold reduction in resistance to β-lactam antibiotics, demonstrating the "β-lactam seesaw effect". Whole-genome sequencing confirmed their isogenic nature (ST59-SCCmecIV-t172), identifying seven mutations in XF2, including those in walK (G223S), vraR (D88Y), clpX (P64L), and ltaS (L62P), as well as a frameshift mutation in mgt (S39fs), likely contributing to resistance. Transmission electron microscopy and autolysis assays demonstrated that XF2 had a thicker cell wall and a slower autolysis rate compared to XF1. Phenotypic analysis showed that XF2 exhibited reduced growth rate, diminished virulence, and enhanced biofilm formation compared to XF1. Gene expression analysis supported these findings, revealing significant alterations in pathways related to cell wall metabolism, autolysis, and virulence regulation. These adaptations highlight the genomic and phenotypic plasticity of MRSA under antibiotic pressure, enabling resistance and persistence. This study underscores the urgent need for enhanced surveillance and alternative therapeutic strategies, including exploiting the β-lactam seesaw effect, to combat lipoglycopeptide-nonsusceptible MRSA.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major global health threat due to its resistance to multiple antibiotics, making conventional treatments ineffective. The rise in antibiotic resistance highlights the urgent need for new therapies. Sortase A (SrtA), a key virulence factor in Staphylococcus aureus (S. aureus), facilitates bacterial adhesion and infection by anchoring surface proteins to host cells, making it a promising drug target. In this study, we investigated the potential of osmundacetone (OSC), a natural compound from Osmundae Rhizoma, as an SrtA inhibitor. Using fluorescence resonance energy transfer (FRET), OSC was found to inhibit SrtA with an IC50 of 1.29 μg/mL (7.24 μM). Further in vitro assays confirmed the effectiveness of OSC in inhibiting SrtA-mediated bacterial adhesion, invasion and biofilm formation. Fluorescence quenching and molecular docking pinpointed the binding site of OSC on SrtA. In vivo, OSC improved survival rates in MRSA-infected mice and Galleria mellonella (G. mellonella) while reducing bacterial loads in infected tissues. These results suggest OSC as a promising candidate for anti-MRSA therapies.

Introduction: The use of extracorporeal membrane oxygenation (ECMO) in children continues to increase nationally, including patients with methicillin-resistant Staphylococcus aureus (MRSA) infection. Survival of pediatric patients with MRSA sepsis has not improved over the last 20 years. We sought to review our institutional experience and outcomes of ECMO support among children with MRSA infection.Methods: Children aged 0-19 years who received ECMO support from October 2014 to June 2021 were reviewed retrospectively. Patients with laboratory confirmed MRSA infections were identified.Results: Out of 88 unique pediatric patients requiring ECMO support, eight patients had documented MRSA infections. The duration of mechanical ventilation prior to ECMO initiation was an average of seven days (range 0.7 to 21.8 days). The median ECMO duration was 648.1 h (range 15.5 to 1580.5 h). Five patients were successfully decannulated; however, only two patients survived to discharge. The two surviving patients were both cannulated via VV-ECMO. Mechanical ventilation prior to ECMO was 4.5 and 21.8 days in these cases with run durations of 18.9 and 29.9 days, respectively.Conclusions: Our institutional survival of patients with MRSA on ECMO is lower than what has been reported in recent database studies, but notably, 62.5% were successfully decannulated. While both surviving patients were supported with VV-ECMO, there was no other clear trend in factors that contributed to survival. MRSA continues to be a source of significant morbidity and mortality among pediatric patients. On-going investigation of outcomes and factors contributing to survival in patients with MRSA infection on ECMO is warranted.

Antimicrobial resistance is a critical public health threat, compromising treatment effectiveness. The spread of resistant pathogens, facilitated by genetic variability and horizontal gene transfer, primarily through plasmids, poses significant challenges to health systems.

This review explores the potential of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology and Cas9 nucleases in combating antimicrobial resistance.

The literature review followed the PRISMA guidelines using PubMed, Embase, and Scopus databases until July 2023.

The Enterobacterales family, particularly Escherichia coli, was the main focus. The resistance genes targeted were mainly associated with β-lactam antibiotics, specifically bla genes, and colistin resistance linked to the mcr-1 gene. Plasmid vectors have been the primary delivery method for the CRISPR-Cas9 system, with conjugative plasmids resensitizing bacterial strains to various antimicrobials. Other delivery methods included electroporation, phage-mediated delivery, and nanoparticles. The efficacy of the CRISPR-Cas9 system in resensitizing bacterial strains ranged from 4.7% to 100%.

Despite challenges in delivery strategies and clinical application, studies integrating nanotechnology present promising approaches to overcome these limitations. This review highlights new perspectives for the clinical use of CRISPR-Cas9 as a specific and efficient antimicrobial agent, potentially replacing traditional broad-spectrum antimicrobials in the future.

This study aimed to investigate the "seesaw effect" of daptomycin (DAP) and ceftobiprole (BPR) on DAP-resistant (DAP-R) methicillin-resistant Staphylococcus aureus (MRSA) isolates.

Broth microdilution minimum inhibitory concentrations (MICs) of DAP and BPR were tested for laboratory-derived and clinical DAP-R MRSA isolates to estimate the "seesaw effect." Time-kill curves for seven representative DAP-R isolates were obtained using DAP and BPR to validate their synergistic activity in vitro. Whole genome sequencing as well as deletion and complementation of the mprF gene were performed to investigate the mechanisms of the "seesaw effect."

The BPR MICs decreased by half-fold in DAP-R MRSA isolates. The synergistic effect of DAP and BPR against representative clinical and community-associated MRSA (CA-MRSA) isolates was demonstrated in time-kill analyses, showing that synergistic activity was preferred in CA-MRSA compared with hospital-associated MRSA. The mprF mutations were identified in isolates exhibiting the "seesaw effect." These mutations increased the DAP MIC while decreasing the BPR MIC.

The "seesaw effect" between DAP and BPR was prevalent among DAP-R MRSA isolates. This phenomenon was associated with the mprF mutations of MRSA.

Animal infection models are essential for understanding bacterial pathogenicity and corresponding host immune responses. In this study, we investigated whether juvenile Xenopus laevis could be used as an infection model for human pathogenic bacteria. Xenopus frogs succumbed to intraperitoneal injection containing the human pathogenic bacteria Staphylococcus aureus, Pseudomonas aeruginosa, and Listeria monocytogenes. In contrast, non-pathogenic bacteria Bacillus subtilis and Escherichia coli did not induce mortality in Xenopus frogs. The administration of appropriate antibiotics suppressed mortality caused by S. aureus and P. aeruginosa. Strains lacking the agr locus, cvfA (rny) gene, or hemolysin genes in S. aureus, LIPI-1-deleted mutant of L. monocytogenes, which attenuate virulence within mammals, exhibited reduced virulence in Xenopus frogs compared with their respective wild-type counterparts. Bacterial distribution analysis revealed that S. aureus persisted in the blood, liver, heart, and muscles of Xenopus frogs until death. These results suggested that intraperitoneal injection of human pathogenic bacteria induces sepsis-like symptoms in Xenopus frogs, supporting their use as a valuable animal model for evaluating antimicrobial efficacy and identifying virulence genes in various human pathogenic bacteria.

Methicillin-resistant Staphylococcus aureus (MRSA), as one of the main pathogens causing skin and soft tissue infections, poses challenges in treatment due to its high resistance to antibiotics. As one of the efficacious essential oil components in numerous traditional Chinese medicines, linalool was believed to possess antimicrobial activity against pathogenic microorganisms. Here, we investigated the therapeutic effects of linalool on MRSA-infected mice by examining their post-treatment outcomes. This was done through observations of physiological conditions, pathological sections, inflammatory factors, and changes in the skin microenvironment. We have confirmed the effectiveness of linalool in treating MRSA infections. Mice treated with linalool exhibited more pronounced signs of recovery, such as reduced skin necrosis, increased fibroplasia, greater neovascularization, and resolution of inflammatory infiltration. In addition, there was an improvement in the inflammatory environment, with a decrease in inflammatory factors. The microbial composition on the skin surface also confirmed this improvement. After linalool treatment, mice exhibited better species diversity on the skin, making it easier to maintain the skin's homeostasis. The excellent performance of linalool in combating MRSA infections provides a new direction for the search for new antibiotics against multidrug-resistant bacteria, highlighting the potential of linalool as a promising anti-MRSA drug.

Intracellular microbes are actively present in various tumor types in low biomass and play a major role in metastasis. Eliminating intracellular microbes on a cellular level with precision remains a challenge. To address this issue, we designed a screening pipeline to characterize intracellular microbes and their interaction with host cells. We used host and microbial in vitro lab-based constant and reproducible model, host as (mammalian cancer HeLa), and microbial strain as (Escherichia coli 25922). To study the pharmacological impact on intracellular bacterial load, we used antibiotics (ampicillin, roxithromycin, and ciprofloxacin) and chemotherapy drugs (doxorubicin and cisplatin) as external stimuli for both host and microbes. We found that increasing pharmacological stress does not increase microbial load inside the host cells. Eliminations of intracellular bacteria was done by using permutation orthogonal arrays (POA), whereby we acquired optimal drug combination in particular sequence of drugs, which reduced 90%-95% of the intracellular microbial load. Proteomic analysis revealed that upon invasion of Escherichia coli 25922, HeLa cells enriched ATP production pathways to activate intermediate filaments, which should be investigated closely via in vivo models.

The objective of this research was to investigate the genomic epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) in a pediatric population in Shanghai, China. Whole-genome sequencing was conducted for 492 randomly selected MRSA isolates obtained from a pediatric hospital between 2013 and 2022. ST59 (37.4%), ST398 (22.4%), ST88 (5.7%), and ST22 (5.5%) were the predominant lineages among these children. While ST59 maintained a dominant annual proportion before 2017, the proportion of ST398 gradually increased from 2013 to 2016, with ST398 ultimately emerging as a prevalent clone with a proportion comparable to that of ST59 after 2017. Among the prevalent STs, the spa-SCCmec structure also experienced dynamic changes. Within ST59, the t437-IV subtype experienced a decline and has even been replaced by t172-IV in recent years. In ST398, the t011-V subtype appeared in 2014 and rapidly became the leading subtype. The antibiotic resistance profiles and virulence factors exhibited clone-related features. Compared with other prevalent lineages, ST59 presented high resistance to erythromycin and clindamycin, whereas ST398 presented relatively low resistance to common antimicrobial agents and fewer virulence determinants. Panton-Valentine leucocidin was more common in ST338 and ST1232, whereas toxic shock syndrome toxin was closely associated with ST1 and ST5. The MRSA cases could also be classified into community- and hospital-associated cases, with highly significant differences between the two in terms of demographic characteristics, clindamycin susceptibility, and virulence genes. In conclusion, this study revealed high genetic diversity and dynamic changes in the molecular epidemiology of pediatric MRSA isolates from Shanghai collected over a decade.

Methicillin-resistant Staphylococcus aureus (MRSA) has emerged as a significant global health concern. Previous research on MRSA epidemiology has predominantly focused on adult populations or targeted specific infection sites, while there was limited research on the long-term evolution of MRSA from the pediatric population. This study addresses this knowledge gap by conducting a comprehensive, 10-year surveillance of pediatric MRSA isolates using whole-genome sequencing. We characterized the molecular typing, as well as the phenotypic and genotypic antimicrobial resistance profiles, and virulence factors present in MRSA isolates obtained from children. Our results highlight the imperative for continuous, vigilant monitoring of MRSA within the pediatric demographic to track its evolving genetic landscape.

The endoplasmic reticulum (ER) stress sensor inositol-requiring enzyme 1α (IRE1α) is associated with lung infections where innate immune cells are drivers for progression and resolution ammatory cytokinesflammation. Yet, the role of IRE1α in pulmonary innate immune host defense during acute respiratory infection remains unexplored. Here, we found that activation of IRE1α in infected lungs compromises immunity against methicillin-resistant Staphylococcus aureus (MRSA)-induced primary and secondary pneumonia. Moreover, activation of IRE1α in MRSA-infected lungs and alveolar macrophages (AMs) leads to exacerbated production of inflammatory mediators followed by cell death. Ablation of myeloid IRE1α or global IRE1α inhibition confers protection against MRSA-induced pneumonia with improved survival, bacterial clearance, cytokine reduction, and lung injury. In addition, loss of myeloid IRE1α protects mice against MRSA-induced secondary to influenza pneumonia by promoting AM survival. Thus, activation of IRE1α is detrimental to pneumonia, and therefore, it shows potential as a target to control excessive unresolved lung inflammation.

Amphibian skin-secreted antimicrobial peptides (AMPs) have garnered significant attention for their excellent biological activity and low propensity for drug resistance over the past 40 years. Bombinins and bombinin H, two classes of AMPs isolated from the skin secretions of Bombina species, demonstrate strong antimicrobial activity against broad-spectrum microorganisms. In this study, two novel peptides, bombinin-like peptide 7S and bombinin-H2L, were identified from the toad, Bombina variegata. While both peptides exhibited broad-spectrum antimicrobial activity, they also showed relatively high cytotoxicity. To explore the structure-activity relationship and enhance therapeutic potential, bombinin-H2L, which displayed stronger average antimicrobial activity, was used as a template. With the aid of bioinformatics analysis, a series of bombinin-H2L analogues were designed by increasing the net positive charges and/or adjusting the amphiphilicity of the parent peptide. Among these analogues, [Arg8, 15]BH2L and [Lys7, 8]BH2L demonstrated high therapeutic efficacy and specificity toward clinically isolated, drug-resistant Staphylococcus aureus strains in both in vitro and ex vivo tests. Their notable biosafety profiles, sensitivity to diverse environments, and ability to disrupt biofilms highlight their potential for further development. Additionally, studies on the mechanism of [Arg8, 15]BH2L and [Lys7, 8]BH2L revealed a membrane-targeted antimicrobial mechanism, with its antibacterial function exerted by disrupting the integrity of bacterial membranes. These findings provide valuable insights into structural modifications of bombinin H peptides for enhanced activity, and [Arg8, 15]BH2L and [Lys7, 8]BH2L have the potential as promising candidates for novel antibacterial agents in treated bacterial skin infections.

The rise in multidrug resistance and strong biofilm-forming ability of Staphylococcus aureus has led to significant public health concerns. Phage or phage-derived components, such as depolymerase or endolysin, have been considered as potential alternatives to antibiotics for combating antibiotic-resistant bacterial infections. In this study, we cloned and expressed a Staphylococcus aureus phage endolysin, LysP4, and identified its lytic activity. The bactericidal effect of LysP4 was more pronounced against planktonic cells in the logarithmic phase compared to those in the stationary phase. LysP4 reduces bacterial counts by 3 log CFU/mL in 60 min and about 2 log CFU/mL during the stationary phase. LysP4 exhibited optimal lytic activity at pH 5.0-7.0 and remained stable across a temperature range of 16 to 40 °C, with maximal activity observed at 37 °C. LysP4 effectively targets 31 of 38 Staphylococcus strains and successfully eliminates biofilms, reducing bacterial counts by 4 log CFU/mL when combined with vancomycin. Notably, LysP4 demonstrated no hemolytic effects on human red blood cells and no toxic effects on embryonic kidney cells or lung cancer cells. Based on these findings, we believe that LysP4 holds promise as a biological control agent against Staphylococcus infections.

Cutaneous leishmaniasis is a parasitic infection that causes a spectrum of pathology ranging from single, self-healing lesions to disfiguring chronic wounds. In severe disease, uncontrolled inflammation exacerbates tissue damage and delays healing, though the contributing factors are unclear. We previously observed that delayed healing was associated with Staphylococcus aureus in the lesional microbiota of patients with cutaneous leishmaniasis. To investigate how S. aureus impacts immunopathology during leishmania infection, we established a murine model of S. aureus colonization with clinical isolates followed by Leishmania infection. S. aureus triggered early production of interleukin (IL)-1β during Leishmania infection, which was critical for neutrophil recruitment and cutaneous inflammation. S. aureus isolates differentially induced IL-1β and neutrophil recruitment, and isolates that induced greater neutrophil recruitment were resistant to neutrophil killing and persisted longer. We reveal a mechanism whereby S. aureus mediates immunopathology during cutaneous leishmaniasis, suggesting IL-1β as a promising immunomodulatory target for non-healing infections.

Gram-positive superbugs resistant to methicillin and vancomycin pose a severe threat to global public health, urgently demanding novel therapeutic strategies. Herein, we rationally designed and synthesized vancomycin derivatives modified with diverse aryl sulfonium moieties to reactivate its antibacterial potency. By optimizing the sulfonium-based SAR, we got derivatives 2-3 orders of magnitude more active in vitro than vancomycin. Subsequently, preliminary toxicity evaluations for the optimal derivative, 7e, indicated a favorable therapeutic index, while pharmacokinetic assays revealed its good properties, suggesting great drug-like potential. Notably, 7e showed extremely potent in vivo protection efficacy by only a single-dose treatment in the challenging methicillin-resistant Staphylococcus aureus and VRE lethal sepsis mice models. Moreover, two independent and synergistic mechanisms of action were uncovered: membrane perturbation and enhanced cell wall biosynthesis inhibition. These findings revealed the unknown role of sulfonium strategy in vitro and in vivo and positioned 7e as a promising candidate for future development.

Surface contamination is an important, if under-discussed, route of infection transmission. In this study, we suspended chlorhexidine digluconate (CHX) in epoxy resin. CHX was found to be stably incorporated into the material, and its addition to epoxy resin was found to have minimal effects on the optical transparency of the material. After application of the epoxy resin to steel surfaces, time-of-flight secondary ion mass spectrometry revealed that CHX was uniformly present over the surface. Surfaces painted with CHX-resin were found to have significant, reproducible antimicrobial efficacy against E. coli, S. aureus, and C. albicans. We have shown that the addition of CHX has minimal effects on the adhesion of the epoxy resin to surfaces, as well as a high durability of the antimicrobial efficacy. We believe that this material has a wide array of applications, and could be utilised to confer significant, low-cost antimicrobial efficacy to existing surfaces, to prevent surface contamination, and to stop the transmission of infectious disease.

The development of antibiotic-independent antimicrobial materials is critical for addressing bacterial resistance to conventional antibiotics. Currently, there is a lack of comprehensive understanding of ionic liquid-modified composites in antimicrobial applications. Here, we innovatively prepared GO-based composites modified with phosphonate ionic liquids via a series of surface functionalizations. The resulting antibacterial composites exhibit significant broad-spectrum activity against both Gram-negative and Gram-positive bacteria, including drug-resistant strains, with stronger efficacy against Gram-negative species. Additionally, the material features excellent long-term reusability and the ability to inhibit/destroy biofilms, which is vital for combating persistent infections. Mechanistic studies reveal its antibacterial effects through multiple pathways: disrupting bacterial membranes, inducing ROS, and inactivating intracellular substances-mechanisms less likely to promote resistance. Overall, these phosphonate ionic liquid-modified polycationic materials demonstrate substantial potential in treating bacterial infections, offering a promising strategy to tackle antibiotic resistance challenges.

Staphylococcus aureus is one of the most prevalent antibiotic-resistant bacteria, characterized by high morbidity and mortality. The pathogenicity of S. aureus relies on the production of multiple virulence factors. In recent years, antivirulence strategies have shown promise in developing antiinfective drugs by targeting the inhibition of bacterial virulence factors rather than directly killing pathogens. In Asia, some traditional Chinese medicines have a long history of antiinfective application and have demonstrated therapeutic efficacy. However, their antiinfective mechanism has not been fully elucidated. Recent studies have revealed that numerous extracts of TCM, as well as pure compounds from TCM, significantly inhibited the expression of virulence factors of S. aureus, which might be one of their antiinfective mechanisms with potential for the development of novel antiinfective agents. In this review, we summarized the major virulence factors of S. aureus and recent advances in TCM-derived antivirulence agents, including TCM formulae, single herbs, and isolated bioactive compounds, which showed antivirulence capability against S. aureus. Investigating the antivirulence mechanism of TCM not only enhances our understanding of TCM's antiinfective mechanisms but also facilitates the isolation of active compounds with therapeutic potential against S. aureus infection.

MXenes, a rapidly emerging class of two-dimensional materials, have demonstrated exceptional versatility and functionality across various domains, including microbiology and virology. Recent advancements in MXene synthesis techniques, encompassing both top-down and bottom-up approaches, have expanded their potential applications in pathogen detection, antimicrobial treatments, and biomedical platforms. This review highlights the unique physicochemical properties of MXenes, including their large surface area, tunable surface chemistry, and high biocompatibility, which contribute to their antimicrobial efficacy against bacteria, fungi, and viruses, such as SARS-CoV-2. The antibacterial mechanisms of MXenes, including membrane disruption, reactive oxygen species (ROS) generation, and photothermal inactivation, are discussed alongside hybridization strategies that enhance their bioactivity. Additionally, the challenges and future prospects of MXenes in developing advanced antimicrobial coatings, diagnostic tools, and therapeutic systems are outlined. By addressing current limitations and exploring innovative solutions, this study underscores the transformative potential of MXenes in microbiology, virology, and biomedical applications.

Staphylococcus aureus is a common bacterium that sometimes causes various pyogenic diseases. Methicillin resistant S. aureus (MRSA) infections are particularly difficult to treat. Recently, MRSA has been spreading in the community, so it is important to determine the prevalence of MRSA in the community and to conduct epidemiological studies from genetic and statistical perspectives. In this study, S. aureus/MRSA was isolated from the oral and nasal cavities of 504 dental patients (65 inpatients and 439 outpatients). Sixty-two S. aureus strains and 9 MRSA strains were isolated from the oral cavity, and 112 S. aureus strains and 21 MRSA strains were isolated from the nasal cavity. Multi-locus sequence typing (MLST) analysis showed ST8 was high (18 isolates) among 30 MRSA isolates, whereas among 144 methicillin sensitive isolates, ST15 (25 isolates) and ST8 (20 isolates) were high. Statistical analysis of the patients' clinical status revealed a correlation between oral S. aureus and denture use. Among the 34 patients from whom S. aureus was isolated from both sites, 25 had the same ST, and 23 showed below 40 single-nucleotide polymorphisms which are considered to be identical strains. Our study revealed various properties of S. aureus/MRSA in the oral and nasal cavities as commensals.

Bacterial genomic mutations in Staphylococcus aureus have been detected in isolated resistant clinical strains, yet their mechanistic effect on the development of antimicrobial resistance remains unclear. Resistance-associated regulatory systems acquire adaptive mutations under stress conditions that may lead to a gain-of-function effect and contribute to the resistance phenotype. Here, we investigate the effect of a single-point mutation (T331I) in VraS histidine kinase, part of the VraSR two-component system in S. aureus. VraSR senses and responds to environmental stress signals by upregulating gene expression for cell wall synthesis. A combination of enzyme kinetics, microbiological, and transcriptomic analyses revealed the mechanistic effect of the mutation on VraS and S. aureus. Michaelis-Menten kinetics show that the VraS mutation caused an increase in the autophosphorylation rate of VraS and enhanced its catalytic efficiency. The introduction of the mutation through recombineering coupled with CRISPR-Cas9 counterselection to the Newman strain wild-type (WT) genome doubled the minimum inhibitory concentration of three cell wall-targeting antibiotics. The mutation caused an enhanced S. aureus growth rate at sub-lethal doses of the antibiotics, confirming the causative effect of the mutation on bacterial persistence. Transcriptomic analysis showed a genome-wide alteration in gene expression levels and protein-protein interaction network of the mutant compared to the WT strain after exposure to vancomycin. The results suggest that the vraS mutation causes several mechanistic changes at the protein and cellular levels that favor bacterial survival under antibiotic stress and cause the mutation-harboring strains to become the dominant population during infection.IMPORTANCERising antimicrobial resistance (AMR) is a global health problem. Mutations in the two-component system have been linked to drug resistance in Staphylococcus aureus, yet the exact mechanism through which these mutations work is understudied. We investigated the T331I mutation in the vraS gene linked to sensing and responding to cell wall stress. The mutation caused changes at the protein level by increasing the catalytic efficiency of VraS kinase activity. Introducing the mutation to the genome of an S. aureus strain resulted in changes in phenotypic antibiotic susceptibility, growth kinetics, and genome-wide transcriptomic alterations. By a combination of enzyme kinetics, microbiological, and transcriptomic approaches, we highlight how small genetic changes can significantly impact bacterial physiology and survival under antibiotic stress. Understanding the mechanistic basis of antibiotic resistance is crucial to guide the development of novel therapeutic agents to combat AMR.

The emergence and dissemination of antibiotic resistance represents a significant and ever-increasing global threat to human, animal, and environmental health. The explosive proliferation of resistance has ultimately been seen in all clinically used antibiotics. Infections caused by antibiotic-resistant bacteria have been associated with an estimated 4,950,000 deaths annually, with extremely limited therapeutic options and only a few new antibiotics under development. To combat this silent pandemic, a better understanding of the molecular mechanisms of antibiotic resistance is immensely needed, which not only helps to improve the efficacy of current drugs in clinical use but also design new antimicrobial agents that are less susceptible to resistance.

The past few years have witnessed a number of new advances in revealing the molecular mechanisms of AMR. Following five sophisticated mechanisms (efflux pump, antibiotics inactivation by enzymes, alteration of membrane permeability, target modification, and target protection), the roles of various novel proteins/enzymes in the acquisition of antibiotic resistance are constantly being described. They are widely used by clinical bacterial strains, playing a key role in the emergence of resistance.

While most of these have so far received less attention, expanding our understanding of these emerging resistance mechanisms is of crucial importance to combat the antibiotic resistance crisis in the world. This review summarizes recent advances in our knowledge of emerging resistance mechanisms in bacteria, providing an update on the current antibiotic resistance threats and encouraging researchers to develop critical strategies for overcoming the resistance.

UDP-N-acetylglucosamine-undecaprenyl-phosphate N-acetylglucosaminephosphotransferase (TarO) has been found to simultaneously contribute to β-lactam resistance and virulence of Methicillin-resistant Staphylococcus aureus (MRSA). However, optimization of hit compounds targeting TarO has been hindered due to their high lipophilicity and the poor correlation between the enzyme activity inhibition and β-lactam sensitization. In this study, 31 analogues of Tarocin A were designed, synthesized and evaluated by a luminescence-based reporter preliminary screening. In the subsequent β-lactams synergy test, a good correlation was observed between the results obtained from these two methods. Finally, analog 18a with more potential against TarO and an improved hydrophilicity (clogP = 3.2) was obtained. Compared with Tarocin A, 18a shows stronger β-lactam sensitizing and anti-biofilm activities in vitro, as well as potent anti-virulence and synergistic potency with imipenem in vivo. These results suggest that TarO is a promising target for combating MRSA, and 18a can serve as a lead molecule.

Pneumonia caused by Staphylococcus aureus infection has consistently been a significant cause of morbidity and mortality worldwide. Extensive research to date indicates that N6-methyladenosine (m6A) modification plays a crucial role in the development and progression of various diseases. However, it remains unknown whether the m6A modification affects the progression of bacterial pneumonia. To explore this question, we assessed the levels of m6A as well as the expression of methyltransferases (METTL3 and METTL14), demethylase fat mass and obesity-related protein (FTO), and methylation reader proteins YTHDF1 and YTHDF2 in mice and MH-S cells during S. aureus infection. The levels of m6A and METTL3 were significantly upregulated in S. aureus-infected mice and MH-S cells. siMETTL3 knockdown resulted in more severe bacterial colonization, lung damage, increased inflammatory cytokines (IL-6, IL-1β, TNF-α), and mortality rates in mice as well as MH-S cells following the bacterial infection. Regulation of lung inflammation levels by METTL3 was associated with the activation of the MAPK/NF-κB/JAK2-STAT3 signaling pathway. Moreover, siMETTL3 mice exhibited an increased release of superoxides and exacerbated oxidative stress in the lungs following S. aureus infection, which was correlated with impaired mitochondrial autophagy mediated by the Pink1/Parkin pathway. Our findings provide previously unrecognized evidence of the protective role of METTL3 in S. aureus-induced acute pneumonia, indicating a potential therapeutic target for S. aureus infections.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of hospital- and community-acquired infections, necessitating the development of novel antibacterials. Here, we designed and synthesized 30 osthole derivatives with pyridinium quaternary ammonium moieties. In vitro bioassay showed that compounds 8u and 8ac exhibited potent antibacterial activity against S. aureus ATCC 29213 and ten clinical MRSA isolates (MIC = 0.5-1 μg/mL), with low hemolytic activity, rapid bactericidal effects, and minimal resistance induction. In MRSA-infected mouse models of skin abscesses and sepsis, 8u and 8ac also displayed excellent antibacterial effects and safety, which were comparable to vancomycin. Mechanistic studies revealed that 8u and 8ac selectively target bacterial membranes via binding to phosphatidylglycerol (PG), increasing intracellular reactive oxygen species (ROS), inducing content leakage, and ultimately causing bacterial death. These findings suggest 8u and 8ac as promising novel lead candidates for anti-MRSA drug development.