Discovery of Gambogic acid as an antibacterial adjuvant against vancomycin-resistant enterococci in vitro and in vivo.

BACKGROUND: The emergence and spread of vancomycin-resistant enterococci (VRE) have posed a significant challenge to clinical treatment, underscoring the need to develop novel strategies. As therapeutic options for VRE are limited, discovering vancomycin enhancer is a feasible way of combating VRE. Gambogic acid (GA) is a natural product derived from the resin of Garcinia hanburyi Hook.f. (Clusiaceae), which possesses antibacterial activity. PURPOSE: This study aimed to investigate the potential of GA as an adjuvant to restore the susceptibility of VRE to vancomycin. METHODS: In vitro antibacterial and synergistic activities were evaluated against vancomycin-susceptible and resistant strains by the broth microdilution method for the Minimal Inhibitory Concentrations (MICs) determination, and checkerboard assay and time-kill curve analysis for synergy evaluation. In vivo study was conducted on a mouse multi-organ infection model. The underlying antibacterial mechanism of GA was also explored. RESULTS: GA showed a potent in vitro activity against all tested strains, with MICs ranging from 2 to 4 µg/ml. The combination of GA and vancomycin exhibited a synergistic effect against 18 out of 23 tested VRE strains, with a median fractional inhibitory concentration index (FICI) of 0.254, and demonstrated a synergistic effect in the time-kill assay. The combination therapy exhibited a significant reduction in tissue bacterial load compared with either compound used alone. GA strongly binds to the ParE subunit of topoisomerase IV, a bacterial type II DNA topoisomerase, and suppresses its activity. CONCLUSIONS: The study suggests that GA has a significant antibacterial activity against enterococci, and sub-MIC concentrations of GA can restore the activity of vancomycin against VRE in vitro and in vivo. These findings indicate that GA has the potential to be a new antibacterial adjuvant to vancomycin in the treatment of infections caused by VRE.

Subplenones A-J: Dimeric Xanthones with Antibacterial Activity from the Endophytic Fungus Subplenodomus sp. CPCC 401465.

Subplenones A-J (1-10), 10 new xanthone dimers, have been isolated and characterized from the endophytic fungus Subplenodomus sp. CPCC 401465, which resides within the Chinese medicinal plant Gentiana straminea. The isolation process was guided by antibacterial assays and molecular-networking-based analyses. The chemical structures of these compounds were elucidated through the interpretation of nuclear magnetic resonance (NMR) and high-resolution electrospray ionization mass spectrometry (HRESIMS) data. Furthermore, the relative configuration of the compounds was determined using NMR and single-crystal X-ray diffraction analyses, and the absolute configuration was established using electronic circular dichroism calculations. All of the isolated compounds exhibited significant inhibitory activity against Gram-positive bacteria. Notably, compounds 1, 5, and 7 displayed remarkable inhibitory activity against methicillin-resistant Staphylococcus aureus (MRSA) ATCC 700698, with a minimum inhibitory concentration (MIC) of 0.25 µg/mL, and against vancomycin-resistant Enterococcus faecium (VRE) ATCC 700221, with MIC values ranging from 0.5 to 1.0 µg/mL.

Quantitative proteomics approach to investigate the antibacterial response of Helicobacter pylori to daphnetin, a traditional Chinese medicine monomer.

Helicobacter pylori is a Gram-negative bacterium related to the development of peptic ulcers and stomach cancer. An increasing number of infected individuals are found to harbor antibiotic-resistant H. pylori, which results in treatment failure. Daphnetin, a traditional Chinese medicine, has a broad spectrum of antibacterial activity without the development of bacterial resistance. However, the antibacterial mechanisms of daphnetin have not been elucidated entirely. To better understand the mechanisms of daphnetin's effect on H. pylori, a label-free quantitative proteomics approach based on an EASY-nLC 1200 system coupled with an Orbitrap Fusion Lumos mass spectrometer was established to investigate the key protein differences between daphnetin- and non-daphnetin-treated H. pylori. Using the criteria of greater than 1.5-fold changes and adjusted p value <0.05, proteins related to metabolism, membrane structure, nucleic acid and protein synthesis, ion binding, H. pylori colonization and infection, stress reaction, flagellar assembly and so on were found to be changed under daphnetin pressure. And the changes of selected proteins in expression level were confirmed by targeted proteomics. These new data provide us a more comprehensive horizon of the proteome changes in H. pylori that occur in response to daphnetin.

Proteomic Changes of Klebsiella pneumoniae in Response to Colistin Treatment and crrB Mutation-Mediated Colistin Resistance.

Polymyxins are increasingly used as the critical last-resort therapeutic options for multidrug-resistant Gram-negative bacteria. Unfortunately, polymyxin resistance has increased gradually over the past few years. Although studies on polymyxin mechanisms are expanding, systemwide analyses of the underlying mechanism for polymyxin resistance and stress response are still lacking. To understand how Klebsiella pneumoniae adapts to colistin (polymyxin E) pressure, we carried out proteomic analysis of a K. pneumoniae strain cultured with different concentrations of colistin. Our results showed that the proteomic responses to colistin treatment in K. pneumoniae involve several pathways, including (i) gluconeogenesis and the tricarboxylic acid (TCA) cycle, (ii) arginine biosynthesis, (iii) porphyrin and chlorophyll metabolism, and (iv) enterobactin biosynthesis. Interestingly, decreased abundances of class A ß-lactamases, including TEM, SHV-11, and SHV-4, were observed in cells treated with colistin. Moreover, we present comprehensive proteome atlases of paired polymyxin-susceptible and -resistant K. pneumoniae strains. The polymyxin-resistant strain Ci, a mutant of K. pneumoniae ATCC BAA 2146, showed a missense mutation in crrB This crrB mutant, which displayed lipid A modification with 4-amino-4-deoxy-l-arabinose (l-Ara4N) and palmitoylation, showed striking increases in the expression of CrrAB, PmrAB, PhoPQ, ArnBCADT, and PagP. We hypothesize that crrB mutations induce elevated expression of the arnBCADTEF operon and pagP via PmrAB and PhoPQ. Moreover, the multidrug efflux pump KexD, which was induced by crrB mutation, also contributed to colistin resistance. Overall, our results demonstrated proteomic responses to colistin treatment and the mechanism of CrrB-mediated colistin resistance, which may offer valuable information on the management of polymyxin resistance.

Methylation of Daptomycin Leading to the Discovery of Kynomycin, a Cyclic Lipodepsipeptide Active against Resistant Pathogens.

Increased usage of daptomycin to treat infections caused by Gram-positive bacterial pathogens has resulted in emergence of resistant mutants. In a search for more effective daptomycin analogues through medicinal chemistry studies, we found that methylation at the nonproteinogenic amino acid kynurenine in daptomycin could result in significant enhancement of antibacterial activity. Termed "kynomycin," this new antibiotic exhibits higher antibacterial activity than daptomycin and is able to eradicate methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) strains, including daptomycin-resistant strains. The improved antimicrobial activity of kynomycin was demonstrated in in vitro time-killing assay, in vivo wax worm model, and different mouse infection models. The increased antibacterial activity, improved pharmacokinetics, and lower cytotoxicity of kynomycin, compared to daptomycin, showed the promise of the future design and development of next-generation daptomycin-based antibiotics.

Design, synthesis, and bioevaluation of a novel class of (E)-4-oxo-crotonamide derivatives as potent antituberculosis agents.

A series of novel (E)-4-oxo-2-crotonamide derivatives were designed and synthesized to find potent antituberculosis agents. All the target compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis H37Rv(MTB). Results reveal that 4-phenyl moiety at part A and short methyl group at part C were found to be favorable. Most of the derivatives displayed promising activity against MTB with MIC ranging from 0.125 to 4 µg/mL. Especially, compound IIIa16 was found to have the best activity with MIC of 0.125 µg/mL against MTB and with MIC in the range of 0.05-0.48 µg/mL against drug-resistant clinical MTB isolates.

Aminoglycoside-based novel probes for bacterial diagnostic and therapeutic applications.

Specific detection of pathogens has long been recognized as a vital strategy in the control of infectious diseases. Two novel theranostic neomycin analogs exhibit efficient targeting, labelling and killing of broad spectrum bacteria while not damaging macrophage-like cells. Furthermore, lipidated probe 2 clearly showed antibacterial activity against methicillin-resistant S. aureus.

Identification of an anti-TB compound targeting the tyrosyl-tRNA synthetase.

OBJECTIVES: Drug-resistant Mycobacterium tuberculosis poses a great threat to human health. Tyrosyl-tRNA synthetase (TyrRS) is one of the aminoacyl tRNA synthetases that catalyse the attachment of amino acids to their cognate tRNAs and are essential for protein synthesis. There are several distinctive differences between bacterial and human TyrRS and therefore it could be a potential target for developing antimicrobial agents. This study aimed to identify a new anti-TB agent targeting M. tuberculosis TyrRS (MtTyrRS). METHODS: We first used Mycobacterium smegmatis for a phenotypic screening of 20 000 compounds. The hit compounds were then screened with MtTyrRS. The interaction between hit compound IMB-T130 and the target protein was analysed by surface plasmon resonance (SPR) assay and molecular docking experiments. The target of IMB-T130 was further confirmed by the overexpression of the target protein. The antibacterial activity of IMB-T130 against various standard and clinical drug-resistant M. tuberculosis strains was evaluated using the microplate Alamar blue assay. RESULTS: Compound IMB-T130 was identified as a hit compound that inhibits the growth of M. smegmatis and the in vitro activity of MtTyrRS. The interaction between IMB-T130 and MtTyrRS was confirmed by SPR assay and molecular docking analysis. The higher MIC for a strain overexpressing the target protein also suggests that MtTyrRS is likely to be the target of IMB-T130. IMB-T130 shows excellent anti-TB activity and low cytotoxicity. CONCLUSIONS: IMB-T130 inhibits the growth of MDR-TB and XDR-TB by targeting MtTyrRS. Because of its low cytotoxicity against mammalian cells, IMB-T130 is a promising new agent against drug-resistant M. tuberculosis.

Inhibitory effects on bacterial growth and beta-ketoacyl-ACP reductase by different species of maple leaf extracts and tannic acid.

It is important to develop new antibiotics aimed at novel targets. The investigation found that the leaf extracts from five maples (Acer platanoides, Acer campestre, Acer rubrum, Acer saccharum and Acer truncatum Bunge collected in Denmark, Canada and China) and their component tannic acid displayed antibacterial ability against 24 standard bacteria strains with the minimum inhibitory concentration of 0.3-8.0 mg/mL. Unlike the standard antibiotic levofloxacin (LFX), these samples inhibited Gram-positive bacteria more effectively than they inhibited Gram-negative bacteria. These samples effectively inhibited two antidrug bacterial strains. The results show that these samples inhibit bacteria by a different mechanism from LFX. These samples potently inhibited b-ketoacyl-ACP reductase (FabG), which is an important enzyme in bacterial fatty acid synthesis. Tannic acid showed the strongest inhibition on FabG with a half inhibition concentration of 0.78 microM (0.81 microg/mL). Furthermore, tannic acid and two maple leaf extracts showed time-dependent irreversible inhibition of FabG. These three samples also exhibited better inhibition on bacteria. It is suggested that FabG is the antibacteria target of maple leaf extracts and tannic acid, and both reversible and irreversible inhibitions of FabG are important for the antibacterial effect.