Huanqin Dai

High-throughput synergy screening identifies microbial metabolites as combination agents for the treatment of fungal infections

The high mortality rate of immunocompromised patients with fungal infections and the limited availability of highly efficacious and safe agents demand the development of new antifungal therapeutics. To rapidly discover such agents, we developed a high-throughput synergy screening (HTSS) strategy for novel microbial natural products. Specifically, a microbial natural product library was screened for hits that synergize the effect of a low dosage of ketoconazole (KTC) that alone shows little detectable fungicidal activity. Through screening of ≈20,000 microbial extracts, 12 hits were identified with broad-spectrum antifungal activity. Seven of them showed little cytotoxicity against human hepatoma cells. Fractionation of the active extracts revealed beauvericin (BEA) as the most potent component, because it dramatically synergized KTC activity against diverse fungal pathogens by a checkerboard assay. Significantly, in our immunocompromised mouse model, combinations of BEA (0.5 mg/kg) and KTC (0.5 mg/kg) prolonged survival of the host infected with Candida parapsilosis and reduced fungal colony counts in animal organs including kidneys, lungs, and brains. Such an effect was not achieved even with the high dose of 50 mg/kg KTC. These data support synergism between BEA and KTC and thereby a prospective strategy for antifungal therapy.

Medium optimization for the production of avermectin B1a by Streptomyces avermitilis 14-12A using response surface methodology

Response surface methodology was employed to optimize the composition of medium for the production of avermectin B1a by Streptomyces avermitilis 14-12A in shaker flask cultivation. Corn starch and yeast extract were found to have significant effects on avermectin B1a production by the Plackett-Burman design. The steepest ascent method was used to access the optimal region of the medium composition, followed by an application of response surface. The analysis revealed that the optimum values of the tested variables were 149.57 g/l corn starch and 8.92 g/l yeast extract. A production of 5128 mg/l, which was in agreement with the prediction, was observed in verification experiment. In comparison to the production of original level (3528 mg/l), 1.45-fold increase had been obtained.

Bioprospecting microbial natural product libraries from the marine environment for drug discovery

Marine microorganisms are fascinating resources due to their production of novel natural products with antimicrobial activities. Increases in both the number of new chemical entities found and the substantiation of indigenous marine actinobacteria present a fundamental difficulty in the future discovery of novel antimicrobials, namely dereplication of those compounds already discovered. This review will share our experience on the taxonomic-based construction of a highly diversified and low redundant marine microbial natural product library for high-throughput antibiotic screening. We anticipate that libraries such as these can drive the drug discovery process now and in the future.

Abyssomicins from the South China Sea deep-sea sediment Verrucosispora sp.: natural thioether michael addition adducts as antitubercular prodrugs

Tuberculosis (TB) is a leading cause of death in the world today, and is exacerbated by the prevalence of multi- (MDR-TB), extensively (XDR-TB), and totally (TDR-TB) drug resistant strains. Despite the threat to human health, existing frontline TB therapeutics remain constrained to a handful of vintage antibiotics prescribed in a combinatorial format to achieve efficacy. The current shortfall in antitubercular drugs demands urgent attention, to develop new antibiotics effective against all strains of tuberculosis.

Trichodermaketones A-D and 7-O-Methylkoninginin D from the Marine Fungus Trichoderma koningii

Five new polyketide derivatives, 7-O-methylkoninginin D (1) and trichodermaketones A-D (2-5), together with four known compounds, koninginins A, D, E, and F, were isolated from the marine-derived fungus Trichoderma koningii. Trichodermaketones A (2) and B (3) are unprecedented polyketides with a bistetrafuran-containing tricyclic skeleton. The chemical structures and absolute configurations of compounds 1-5 were elucidated by comparing with literature data and extensive spectroscopic methods, including 2D NMR and CD spectroscopic analysis. Compounds 1-5 were evaluated for action against bacteria and fungi and for synergistic antifungal activity. Compound 2 showed synergistic antifungal activity against Candida albicans with 0.05 microg/mL ketoconazole.

Isolation and Structural Elucidation of Proline-Containing Cyclopentapeptides from an Endolichenic Xylaria sp.

Two new cyclic pentapeptides (1 and 2) and the known blazein (3), ganodesterone (4), ergosterin (5), cerevisterol (6), 24-methylcholesta-4,6,8(14),22-tetraen-3-one (7), 5,8-epidioxyergosta-6,22-dien-3-ol (8), 16-α-d-mannopyranosyloxyisopimar-7-en-19-oic acid (9), and 16-hydroxy isopimar-7-en-19-oic acid (10) have been isolated from the crude extract of an endolichenic Xylaria sp. The structures of 1 and 2 were elucidated primarily by NMR and MS methods. The absolute configurations of 1 and 2 were assigned using Marfeys method on their acid hydrolysate. Compounds 1-10 were evaluated for activity against fungi and for synergistic antifungal activity. Compound 1 showed synergistic antifungal activity against Candida albicans SC5314 with 0.004 μg/mL ketoconazole.

Amycolatopsis marina sp nov., an actinomycete isolated from an ocean sediment

A Gram-positive, aerobic, non-motile actinobacterium, designated strain Ms392A(T), was isolated from an ocean-sediment sample collected from the South China Sea. The isolate contained chemical markers that supported chemotaxonomic assignment to the genus Amycolatopsis. On the basis of an analysis of 16S rRNA gene sequence similarities, strain Ms392A(T) represents a novel subclade within the genus Amycolatopsis, with Amycolatopsis palatopharyngis 1BDZ(T) as its closest phylogenetic neighbour (99.4 % similarity). However, DNA-DNA hybridization demonstrated that strain Ms392A(T) was distinct from A. palatopharyngis AS 4.1729(T) (48.6 % relatedness). The polyphasic analysis demonstrated that the ocean isolate can be clearly distinguished from recognized species of the genus Amycolatopsis. Therefore, strain Ms392A(T) represents a novel species of the genus Amycolatopsis, for which the name Amycolatopsis marina sp. nov. is proposed. The type strain is Ms392A(T) (=CGMCC 4.3568(T) =NBRC 104263(T)).

Bioprospecting for antituberculosis leads from microbial metabolites

Microbial metabolites have been an important source of tuberculosis (TB) therapeutics, but the last truly novel drug that was approved for the treatment of TB was discovered 40 years ago. In light of the growing threat of multi-drug resistance, recent advances have been made to accelerate the discovery rate of novel TB drugs including diversifying strategies for environmental strains, and high-throughput screening assays. This review will discuss the approaches used in biodiversity- and taxonomy-guided microbial natural product library construction, specific cell-based and target-based high-throughput screening assays and early-stage dereplication processes by liquid chromatography-mass spectrometry (LC-MS). New antituberculosis natural products that have been recently discovered are highlighted.

Optimization for the Production of Surfactin with a New Synergistic Antifungal Activity

Background Two of our long term efforts are to discover compounds with synergistic antifungal activity from metabolites of marine derived microbes and to optimize the production of the interesting compounds produced by microorganisms. In this respect, new applications or mechanisms of already known compounds with a high production yield could be continually identified. Surfactin is a well-known lipopeptide biosurfactant with a broad spectrum of antimicrobial and antiviral activity; however, there is less knowledge on surfactin’s antifungal activity. In this study, we investigated the synergistic antifungal activity of C15-surfactin and the optimization of its production by the response surface method. Methodology/Principal Findings Using a synergistic antifungal screening model, we found that the combination of C15-surfactin and ketoconazole (KTC) showed synergistic antifungal effect on Candida albicans SC5314 when the concentrations of C15-surfactin and KTC were 6.25 µg/mL and 0.004 µg/mL, respectively. These concentrations were lower than their own efficient antifungal concentrations, which are >100 µg/mL and 0.016 µg/mL, respectively. The production of C15-surfactin from Bacillus amyloliquefaciens was optimized by the response surface methodology in shaker flask cultivation. The Plackett-Burman design found sucrose, ammonium nitrate and NaH2PO4.2H2O to have significant effects on C15-surfactin production. The optimum values of the tested variables were 21.17 g/L sucrose, 2.50 g/L ammonium nitrate and 11.56 g/L NaH2PO4·2H2O. A production of 134.2 mg/L, which were in agreement with the prediction, was observed in a verification experiment. In comparison to the production of original level (88.6 mg/L), a 1.52-fold increase had been obtained. Conclusion/Significance This work first found that C15-surfactin was an efficient synergistic antifungal agent, and demonstrated that response surface methodology was an effective method to improve the production of C15-surfactin.

Synergistic combinations of antifungals and anti-virulence agents to fight against Candida albicans

Candida albicans, one of the pathogenic Candida species, causes high mortality rate in immunocompromised and high-risk surgical patients. In the last decade, only one new class of antifungal drug echinocandin was applied. The increased therapy failures, such as the one caused by multi-drug resistance, demand innovative strategies for new effective antifungal drugs. Synergistic combinations of antifungals and anti-virulence agents highlight the pragmatic strategy to reduce the development of drug resistant and potentially repurpose known antifungals, which bypass the costly and time-consuming pipeline of new drug development. Anti-virulence and synergistic combination provide new options for antifungal drug discovery by counteracting the difficulty or failure of traditional therapy for fungal infections.