Multi-drug-resistant Staphylococcus aureus infections necessitate novel antibiotic development. D-3263, a transient receptor potential melastatin member 8 (TRPM8) agonist, has potential antineoplastic properties. Here, we reported the antibacterial and antibiofilm activities of D-3263. Minimum inhibitory concentrations (MICs) against S. aureus, Enterococcus faecalis and E. faecium were ≤ 50 µM. D-3263 exhibited bactericidal effects against clinical methicillin-resistant S. aureus (MRSA) and E. faecalis strains at 4× MIC. Subinhibitory D-3263 concentrations effectively inhibited S. aureus and E. faecalis biofilms, with higher concentrations also clearing mature biofilms. Proteomic analysis revealed differential expression of 29 proteins under 1/2 × MIC D-3263, influencing amino acid biosynthesis and carbohydrate metabolism. Additionally, D-3263 enhanced membrane permeability of S. aureus and E. faecalis. Bacterial membrane phospholipids phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and cardiolipin (CL) dose-dependently increased D-3263 MICs. Overall, our data suggested that D-3263 exhibited potent antibacterial and antibiofilm activities against S. aureus by targeting the cell membrane.

The overuse of antibiotics in animal farming and aquaculture has led to multidrug-resistant methicillin-sensitive Staphylococcus aureus (MR-MSSA) becoming a common pathogen in foodborne diseases. Sophora flavescens Ait. serves as a traditional plant antibacterial agent and functional food ingredient. A total of 30 compounds (1-30) were isolated from the root bark of S. flavescens, consisting of 20 new compounds (1-20). In the biological activity assay, compound 1 demonstrated a remarkable inhibitory effect on MR-MSSA, with an MIC of 2 μg/mL. Furthermore, 1 was found to rapidly eliminate bacteria, inhibit biofilm growth, and exhibit exceptionally low cytotoxicity. Mechanistic studies have revealed that 1 possesses an enhanced membrane-targeting ability, binding to the bacterial cell membrane components phosphatidylglycerol (PG), phosphatidylethanolamine (PE), and cardiolipin (CL). This disruption of bacterial cell membrane integrity increases intracellular reactive oxygen species, protein and DNA leakage, reduced bacterial metabolism, and ultimately bacterial death. In summary, these findings suggest that compound 1 holds promise as a lead compound against MR-MSSA.

The ubiquitous presence of antimicrobial-resistant organisms and antimicrobial resistance genes (ARGs) constitutes a major threat to global public safety. Tetracycline (TET) is a common antimicrobial agent that inhibits bacterial growth and is frequently detected in aquatic environments. Although TET may display coselection for resistance, limited knowledge is available on whether and how it might influence plasmid-mediated conjugation. Subinhibitory concentrations (3.9-250 ng/mL) of TET promoted horizontal gene transfer (HGT) via the mobilizable plasmid pVP52-1 from the donor Vibrio parahaemolyticus NJIFDCVp52 to the recipient Escherichia coli EC600 by 1.47- to 3.19-fold. The transcription levels of tetracycline resistance genes [tetA, tetR(A)], conjugation-related genes (traA, traD), outer membrane protein genes (ompA, ompK, ompV), reactive oxygen species (ROS)-related genes (oxyR, rpoS), autoinducer-2 (AI-2) synthesis gene (luxS), and SOS-related genes (lexA, recA) in the donor and recipient were significantly increased. Furthermore, the overproduced intracellular ROS generation and increased cell membrane permeability under TET exposure stimulated the conjugative transfer of ARGs. Overall, this study provides important insights into the contributions of TET to the spread of antimicrobial resistance.

BACKGROUND: Peanut (Arachis hypogaea), a vital oil and food crop globally, is susceptible to web blotch which is a significant foliar disease caused by Phoma arachidicola Marasas Pauer&Boerema leading to substantial yield losses in peanut production. Calcium treatment has been found to enhance plant resistance against pathogens.
RESULTS: This study investigates the impact of exogenous calcium on peanut resistance to web blotch and explores its mechanisms. Greenhouse experiments revealed that exogenous calcium treatment effectively enhanced resistance to peanut web blotch. Specifically, amino acid calcium and sugar alcohol calcium solutions demonstrated the best induced resistance effects, achieving reduction rates of 61.54% and 60% in Baisha1016, and 53.94% and 50% in Luhua11, respectively. All exogenous calcium treatments reduced malondialdehyde (MDA) and relative electrical conductivity (REC) levels in peanut leaves, mitigating pathogen-induced cell membrane damage. Exogenous calcium supplementation led to elevated hydrogen peroxide (H2O2) content and superoxide anion (O2∙-) production in peanut leaves, facilitating the accumulation of reactive oxygen species (ROS) crucial for plant defense responses. Amino acid calcium and sugar alcohol calcium treatments significantly boosted activities of peroxidase (POD), superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) in peanut leaves. Activation of these antioxidant enzymes effectively scavenged excess ROS, maintaining ROS balance and mitigating cellular damage.
CONCLUSIONS: In summary, exogenous calcium treatment triggered ROS production, which was subsequently eliminated by the activation of antioxidant enzymes, thereby reducing cell membrane damage and inducing defense responses against peanut web blotch.

The utilization of natural products in food preservation represents a promising strategy for the dual benefits of controlling foodborne pathogens and enhancing the nutritional properties of foods. Among the phytonutrients, flavonoids have been shown to exert antibacterial effects by disrupting bacterial cell membrane functionality; however, the underlying molecular mechanisms remain elusive. In this study, we investigated the effect of quercetin on the cell membrane permeability of Staphylococcus aureus ATCC 27217. A combined metabolomic and transcriptomic approach was adopted to examine the regulatory mechanism of quercetin with respect to the fatty acid composition and associated genes. Kinetic analysis and molecular docking simulations were conducted to assess quercetin\'s inhibition of β-ketoacyl-acyl carrier protein reductase (FabG), a potential target in the bacterial fatty acid biosynthesis pathway. Metabolomic and transcriptomic results showed that quercetin increased the ratio of unsaturated to saturated fatty acids and the levels of membrane phospholipids. The bacteria reacted to quercetin-induced stress by attempting to enhance fatty acid biosynthesis; however, quercetin directly inhibited FabG activity, thereby disrupting bacterial fatty acid biosynthesis. These findings provide new insights into the mechanism of quercetin\'s effects on bacterial cell membranes and suggest potential applications for quercetin in bacterial inhibition.

The Escherichia coli (E.coli) degrading glucose irradiated by ultrasound irradiation (20 W, 14 min) was investigated as the model system, the glucose degradation increased by 13 % while the E.coli proliferation decreased by 10 % after culture for 18 h. It indicated a tradeoff effect between substrate degradation and cell proliferation, which drove the enhanced contaminants removal and excess sludge reduction in a weak ultrasound enhanced biological wastewater treatment. The enzymatic activities (catalase, superoxide dismutase, adenosine triphosphatases, lactic dehydrogenase, membrane permeability, intracellular reactive oxygen species and calcium ion of E. coli increased immediately by 12 %, 63 %, 124 %, 19 %, 15 %, 4-fold and 38-fold, respectively by ultrasound irradiation power of 20 W for 14 min. Furthermore, the membrane permeability of irradiated E. coli increased by 26 % even though the ultrasound stopped for 10 h. Additionally, pathways associated with glucose degradation and cell proliferation were continuously up-regulated and down-regulated, respectively.

A myriad of nonantibiotic compounds is released into the environment, some of which may contribute to the dissemination of antimicrobial resistance by stimulating conjugation. Here, we analyzed a collection of studies to (i) identify patterns of transfer stimulation across groups and concentrations of chemicals, (ii) evaluate the strength of evidence for the proposed mechanisms behind conjugal stimulation, and (iii) examine the plausibility of alternative mechanisms. We show that stimulatory nonantibiotic compounds act at concentrations from 1/1000 to 1/10 of the minimal inhibitory concentration for the donor strain but that stimulation is always modest (less than 8-fold). The main proposed mechanisms for stimulation via the reactive oxygen species/SOS cascade and/or an increase in cell membrane permeability are not unequivocally supported by the literature. However, we identify the reactive oxygen species/SOS cascade as the most likely mechanism. This remains to be confirmed by firm molecular evidence. Such evidence and more standardized and high-throughput conjugation assays are needed to create technologies and solutions to limit the stimulation of conjugal gene transfer and contribute to mitigating global antibiotic resistance.

Potyviridae, the largest family of plant RNA viruses, includes many important pathogens that significantly reduce the yields of many crops worldwide. In this study, we report that the 6-kilodalton peptide 1 (6K1), one of the least characterized potyviral proteins, is an endoplasmic reticulum-localized protein. AI-assisted structure modeling and biochemical assays suggest that 6K1 forms pentamers with a central hydrophobic tunnel, can increase the cell membrane permeability of Escherichia coli and Nicotiana benthamiana, and can conduct potassium in Saccharomyces cerevisiae. An infectivity assay showed that viral proliferation is inhibited by mutations that affect 6K1 multimerization. Moreover, the 6K1 or its homologous 7K proteins from other viruses of the Potyviridae family also have the ability to increase cell membrane permeability and transmembrane potassium conductance. Taken together, these data reveal that 6K1 and its homologous 7K proteins function as viroporins in viral infected cells.

Listeria monocytogenes exhibits varying levels of pathogenicity when entering the host through contaminated food. However, little is known regarding the stress response and environmental tolerance mechanism of different virulence strains to host gastrointestinal (GI) stimuli. This study analyzed the differences in the survival and genes of stress responses among two strains of L. monocytogenes 10403S (serotype 1/2a, highly virulent strain) and M7 (serotype 4a, low-virulence strain) during simulated gastrointestinal digestion. The results indicated that L. monocytogenes 10403S showed greater acid and bile salt tolerance than L. monocytogenes M7, with higher survival rates and less cell deformation and cell membrane permeability during the in vitro digestion. KEGG analysis of the transcriptomes indicated that L. monocytogenes 10403S displayed significant activity in amino acid metabolism, such as glutamate and arginine, associated with acid tolerance. Additionally, L. monocytogenes 10403S demonstrated a higher efficacy in promoting activities that preserve bacterial cell membrane integrity and facilitate flagellar protein synthesis. These findings will contribute valuable practical insights into the tolerance distinctions among different virulence strains of L. monocytogenes in the GI environment.

The high prevalence of antibiotic-resistant bacteria poses a threat to the global public health. The appropriate use of adjuvants to restore the antimicrobial activity of antibiotics against resistant bacteria could be an effective strategy for combating antibiotic resistance. In this study, we investigated the counteraction of Triton X-100 (TX-100) and the mechanisms underlying the antibiotic resistance of Enterococcus faecalis (E. faecalis).
Standard, wild-type (WT), and induced antibiotic-resistant E. faecalis strains were used in this study. In vitro antibacterial experiments were conducted to evaluate the antimicrobial activities of gentamicin sulfate and ciprofloxacin hydrochloride in the presence and absence of 0.02 % TX-100 against both planktonic and biofilm bacteria. Transcriptomic and untargeted metabolomic analyses were performed to explore the molecular mechanisms of TX-100 as an antibiotic adjuvant. Additionally, membrane permeability, membrane potential, glycolysis-related enzyme activity, intracellular adenosine triphosphate (ATP), and expression levels of virulence genes were assessed. The biocompatibility of different drug combinations was also evaluated.
A substantially low TX-100 concentration improved the antimicrobial effects of gentamicin sulfate or ciprofloxacin hydrochloride against antibiotic-resistant E. faecalis. Mechanistic studies demonstrated that TX-100 increased cell membrane permeability and dissipated membrane potential. Moreover, antibiotic resistance and pathogenicity of E. faecalis were attenuated by TX-100 via downregulation of the ABC transporter, phosphotransferase system (PTS), and ATP supply.
TX-100 enhanced the antimicrobial activity of gentamicin sulfate and ciprofloxacin hydrochloride at a low concentration by improving antibiotic susceptibility and attenuating antibiotic resistance and pathogenicity of E. faecalis.
These findings provide a theoretical basis for developing new root canal disinfectants that can reduce antibiotic resistance.