Multidrug‐resistant Staphylococcus aureus infections place a huge burden on the healthcare sector and the wider community. An increasing rate of infections caused by methicillin‐resistant Staphylococcus aureus (MRSA) has necessitated the development of alternative agents. We previously reported that usnic acid (UA) has activity against MRSA; here, we report the effect of UA in combination with norfloxacin on the drug resistance of MRSA clinical isolates. We observed that the combination of UA–norfloxacin significantly reduces the bacterial burden in mouse models infected with S. aureus, without causing any detectable associated toxicity. Proteomic analysis indicated that UA–norfloxacin induces oxidative stress within cells, which leads to membrane damage and inhibits metabolic activity and biosynthesis of peptidoglycan and fatty acids. Collectively, this study provides evidence that UA in combination with norfloxacin may be a potential candidate for development into a resistance‐modifying agent for the treatment of invasive MRSA infections.

Curcumin is a natural dietary compound with antimicrobial activity against various gram positive and negative bacteria. This study aims to investigate the proteome level alterations in Bacillus subtilis due to curcumin treatment and identification of its molecular/cellular targets to understand the mechanism of action. We have performed a comprehensive proteomic analysis of B. subtilis AH75 strain at different time intervals of curcumin treatment (20, 60 and 120 min after the drug exposure, three replicates) to compare the protein expression profiles using two complementary quantitative proteomic techniques, 2D-DIGE and iTRAQ. To the best of our knowledge, this is the first comprehensive longitudinal investigation describing the effect of curcumin treatment on B. subtilis proteome. The proteomics analysis revealed several interesting targets such UDP-N-acetylglucosamine 1-carboxyvinyltransferase 1, putative septation protein SpoVG and ATP-dependent Clp protease proteolytic subunit. Further, in silico pathway analysis using DAVID and KOBAS has revealed modulation of pathways related to the fatty acid metabolism and cell wall synthesis, which are crucial for cell viability. Our findings revealed that curcumin treatment lead to inhibition of the cell wall and fatty acid synthesis in addition to differential expression of many crucial proteins involved in modulation of bacterial metabolism. Findings obtained from proteomics analysis were further validated using 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) assay for respiratory activity, resazurin assay for metabolic activity and membrane integrity assay by potassium and inorganic phosphate leakage measurement. The gene expression analysis of selected cell wall biosynthesis enzymes has strengthened the proteomics findings and indicated the major effect of curcumin on cell division.

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Ageratum conyzoides L. (Asteraceae) possesses wound healing, antimicrobial, and anti‐inflammatory activities. Traditionally, this plant is used in skincare. The essential oil (EO) of A. conyzoides aerial parts contains the chromenes precocene I (1) and precocene II (2) as major constituents. This study evaluated precocene II (2) for its in vivo efficacy, mechanistic analysis, and synergistic interaction with norfloxacin against methicillin‐resistant Staphylococcus aureus (MRSA). The study showed that 2 interacted synergistically (Fractional Inhibitory Concentration Index, FICI≤0.5) with norfloxacin and oxacillin and reduced their MIC values significantly. These observations were further validated using the mice model and showed a significant reduction in bacterial load at much lower doses of 2 and norfloxacin without toxicity at 200 mg/kg body weight. Mechanistic studies revealed that 2 regulates bacterial resistance against clinical isolates of S. aureus through membrane disturbance in a dose‐ and time‐dependent manner. Further, precocene II (2) damaged the membrane of the bacteria, as observed from the increased membrane depolarization and uptake of propidium iodide. It also displayed high selectivity towards S. aureus over mammalian cell lines. The in vitro results highlighted the synergistic activity of 2 through the cell membrane damage without any detrimental effect on mammalian cells. In vivo results showed that 2 in combination with norfloxacin significantly reduced bacterial load at much lower concentrations through synergistic interaction without apparent toxicity.