Proteome Analyses of Cellular Proteins in Methicillin-Resistant Staphylococcus aureus Treated with Rhodomyrtone, a Novel Antibiotic Candidate

The ethanolic extract from Rhodomyrtus tomentosa leaf exhibited good antibacterial activities against both methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus ATCC 29213. Its minimal inhibitory concentration (MIC) values ranged from 31.25–62.5 µg/ml, and the minimal bactericidal concentration (MBC) was 250 µg/ml. Rhodomyrtone, an acylphloroglucinol derivative, was 62.5–125 times more potent at inhibiting the bacteria than the ethanolic extract, the MIC and MBC values were 0.5 µg/ml and 2 µg/ml, respectively. To provide insights into antibacterial mechanisms involved, the effects of rhodomyrtone on cellular protein expression of MRSA have been investigated using proteomic approaches. Proteome analyses revealed that rhodomyrtone at subinhibitory concentration (0.174 µg/ml) affected the expression of several major functional classes of whole cell proteins in MRSA. The identified proteins involve in cell wall biosynthesis and cell division, protein degradation, stress response and oxidative stress, cell surface antigen and virulence factor, and various metabolic pathways such as amino acid, carbohydrate, energy, lipid, and nucleotide metabolism. Transmission electron micrographs confirmed the effects of rhodomyrtone on morphological and ultrastructural alterations in the treated bacterial cells. Biological processes in cell wall biosynthesis and cell division were interrupted. Prominent changes including alterations in cell wall, abnormal septum formation, cellular disintegration, and cell lysis were observed. Unusual size and shape of staphylococcal cells were obviously noted in the treated MRSA. These pioneer findings on proteomic profiling and phenotypic features of rhodomyrtone-treated MRSA may resolve its antimicrobial mechanisms which could lead to the development of a new effective regimen for the treatment of MRSA infections

การศึกษาฤทธิ์ของ rhodomyrtone ต่อเซลล์แบคทีเรีย methicillinresistant Staphylococcus aureus โดยใช้เทคนิคทางโปรตีโอมิกส์และทรานสคริปโตมิกส์

Thesis (Ph.D., Microbiology)--Prince of Songkla University, 201

Anti-inflammatory effect of tamarind seed coat extract against LPS-induced RAW264.7 macrophages

Inflammatory response is modulated by stimulated immune cells, and has a pivotal role in host defense system against various stimuli. In this study, we evaluated the anti-inflammatory property of tamarind seed coat extract (TSCE) in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. Various concentrations of TSCE (10, 25, and 50 µg mL-1) were applied and then stimulated with LPS (1 μg mL-1) in RAW264.7 macrophages and the level of reactive oxygen species (ROS) and nitric oxide (NO) were measured. Besides, enzyme-linked immunosorbent assay (ELISA) was used to measure the level of pro-inflammatory cytokines. Our results showed that TSCE suppressed LPS-induced intracellular ROS production and suppressed the NO levels in a dose-dependent manner. Significantly, the anti-inflammatory activity was correlated with a lowered LPS-stimulated TNF-α and IL-1β pro-inflammatory cytokines. These results implied that TSCE possess potent anti-inflammatory activity, which supported new insights into the TSCE utilization to protect inflammation-related disorders

Two dimensional electrophoresis of the of cellular methicillin-resistant <i>S. aureus</i> proteomes.

<p>Protein extracts were prepared from rhodomyrtone-treated cells (A) and untreated control cells (B).</p

Scanning electron micrographs demonstrate the effect of rhodomyrtone on EMRSA-16 cell morphology.

<p>The bacteria were grown in MHB containing 0.5 µg/ml rhodomyrtone (0.5MIC) and incubated for 4 h (A). Untreated control cultures were grown in MHB supplemented with DMSO and incubated for 4 h (B). Scale bars = 2 µm.</p

Schematic representation of the diaminopimelate (DAP) pathway in staphylococci.

<p>The pathway depicts expression of DAP biosynthesis related genes induced or reduced in rhodomyrtone-treated MRSA after 1 h incubation. Gray boxes: genes that were significantly represented by microarray; pink boxes: genes that were significantly represented by qRT-PCR. Blue arrow and red arrow represented an increase or decrease in expression fold change, respectively.</p