The effectiveness of anti-biofilm and anti-virulence properties of dihydrocelastrol and dihydrocelastryl diacetate in fighting against methicillin-resistant Staphylococcus aureus.

Human pathogens have readily been converted into multidrug-resistant pathogens, such as methicillin-resistant Staphylococcus aureus (MRSA), because of the long-term use of conventional antibiotics. In addition, the biofilms formed by S. aureus cells are especially problematic and are related to the persistence of chronic infections because they constitute a major mechanism of promoting tolerance to diverse antimicrobial agents. Hence, the inhibitions of biofilm formation and/or toxin production are accepted as alternative means of controlling S. aureus infections. The present study was aimed at identifying novel anti-biofilm and/or anti-virulence compounds in friedelane-based pentacyclic triterpenoids present in many edible and medicinal plants-and investigating them against MRSA strains. As a result, dihydrocelastrol and dihydrocelastryl diacetate were found to both inhibit the biofilm formation of, and to disrupt the preformed biofilms of, MRSA strains to an increasingly greater degree with increasing concentrations of each compound. Furthermore, these two triterpenoids also clearly inhibited the hemolytic activity of MRSA-and in-line with their anti-biofilm activities, rendered the cell more hydrophilic. Additionally, corroborating phenotypic results, transcriptional analyses showed that both dihydrocelastrol and dihydrocelastryl diacetate disturbed the expression of gene related to α-hemolysin (hla) and down-regulated the expressions of the crucial biofilm-associated genes (agrA, sarA, ica, RNAIII, and rbf) in MRSA. The findings of this study suggest that friedelane-based pentacyclic triterpenoids-especially dihydrocelastrol and dihydrocelastryl diacetate-have the potential to be candidates both for use in controlling biofilm-related infections and for use as important components of anti-virulence strategies for fighting against MRSA infection.

Microbiota Composition and Pulmonary Surfactant Protein Expression as Markers of Death by Drowning.

Pathological diagnosis of drowning remains a challenge for forensic science, because of a lack of pathognomonic findings. We analyzed microbiota and surfactant protein in the lungs for a novel diagnosis of drowning. All rats were divided into drowning, postmortem submersion, and control groups. The water, lungs, closed organs (kidney and liver), and cardiac blood in rats were assayed by targeting 16S ribosomal RNA of Miseq sequencing. Lung samples were analyzed by immunohistochemical staining for surfactant protein A. The closed organs and cardiac blood of drowned group have a lot of aquatic microbes, which have not been detected in postmortem submersion group. Furthermore, intra-alveolar granular staining of surfactant protein A (SP-A) was severely observed in the drowned group than the postmortem submersion and control groups. The findings suggested that the presence of aquatic microbiota in the closed organs and increased expression of SP-A could be markers for a diagnosis of drowning.

Activity of novel inhibitors of Staphylococcus aureus biofilms.

Staphylococcus aureus is one of the most important pathogens causing chronic biofilm infections. These are becoming more difficult to treat owing to drug resistance, particularly because S. aureus biofilms limit the efficacy of antimicrobial agents, leading to high morbidity and mortality. In the present study, we screened for inhibitors of S. aureus biofilm formation using a natural product library from the Korea Chemical Bank (KCB). Screening by crystal violet-based biomass staining assay identified hit compounds. Further examination of antibiofilm properties of these compounds was conducted and led to the identification of celastrol and telithromycin. In vitro, both celastrol and telithromycin were toxic to planktonic S. aureus and also active against a clinical methicillin-resistant S. aureus (MRSA) isolate. The effect of the compounds on preformed biofilms of clinical MRSA isolates was evaluated by confocal laser scanning microscopy (CLSM), which revealed the absence of typical biofilm architecture. In addition, celastrol and telithromycin inhibited the production of extracellular protein at selected sub-MIC concentrations, which revealed the reduced extracellular polymeric substance (EPS) secretion. Celastrol exhibited greater cytotoxicity than telithromycin. These data suggest that the hit compounds, especially telithromycin, could be considered novel inhibitors of S. aureus biofilm. Although the mechanisms of the effects on S. aureus biofilms are not fully understood, our data suggest that telithromycin could be a useful adjuvant therapeutic agent for S. aureus biofilm-related infections.