Discovery and structural assignment of (S)-sydosine from amphipod-derived Aspergillus sydowii MBC15-11F through HRMS, advanced Mosher, and molecular modelling analyses.

AIMS: This study aims to prioritize fungal strains recovered from under-explored habitats that produce new metabolites. HRMS dereplication is used to avoid structure redundancy, and molecular modelling is used to assign absolute configuration. METHODS AND RESULTS: MBC15-11F was isolated from an amphipod and identified using ITS, 28S, and ß-tubulin phylogeny as Aspergillus sydowii. Chemical profiling using taxonomic-based dereplication identified structurally diverse metabolites, including unreported ones. Large-scale fermentation led to the discovery of a new N-acyl adenosine derivative: (S)-sydosine (1) which was elucidated by NMR and HRESIMS analyses. Two known compounds were also identified as predicted by the initial dereplication process. Due to scarcity of 1, molecular modelling was used to assign its absolute configuration without hydrolysis, and is supported by advanced Mosher derivatization. When the isolated compounds were assessed against a panel of bacterial pathogens, only phenamide (3) showed anti-Staphylococcus aureus activity. CONCLUSION: Fermentation of A. sydowii yielded a new (S)-sydosine and known metabolites as predicted by HRESIMS-aided dereplication. Molecular modelling prediction of the absolute configuration of 1 agreed with advanced Mosher analysis.

Antiproliferative potential of moluccella laevis L. aerial parts family lamiaceae (labiatae), supported by phytochemical investigation and molecular docking study.

The current biologically guided study aimed the in vitro investigation of cytotoxic activity, identification of the phytochemical content of Moluccella laevis L. aerial parts and supporting this activity by a molecular docking study. Aqueous fraction demonstrated the most potent cytotoxic effect against CACO-2 with IC50 = 0.067 ± 0.01 µg/mL. Furthermore, EtOAc fraction showed a remarkable cytotoxic activity against MCF-7 cell line with IC50 = 0.35 ± 0.02 µg/mL. Consequently, total ethanolic extract (TEE) and its fractions were subjected to LC-HR-ESI-MS metabolic profiling to discover the constituents that possibly underlie their cytotoxicity. Twenty compounds were tentatively identified from metabolic analysis. Furthermore, eight compounds were isolated. In silico docking study revealed that stachydrine is more likely to account for the antiproliferative activity of both EtOAc and aqueous fractions, probably via its moderate inhibition of receptor tyrosine kinases. [Formula: see text].

Potent antiplasmodial alkaloids from the rhizobacterium Pantoea agglomerans as hemozoin modulators.

Global health concern regarding malaria has increased since the first report of artemisinin-resistant Plasmodium falciparum (Pf) two decades ago. The current therapies suffer various drawbacks such as low efficacy and significant side effects, alarming for an urgent need of more effective and less toxic drugs with higher patient compliance. Chemical entities with natural origins become progressively attractive as new drug leads due to their structural diversity and bio-compatibility. This study initially aimed at the targeted isolation of hydroxyquinoline derivatives following our published genomics and metabolomics study of Pantoea agglomerans (Pa). Fermentation of Pa on a pre-selected medium followed by chromatographic isolation, NMR and HRMS analyses led to the characterisation of one new hydroxyquinoline alkaloid together with another six known congeners and two known hydroxyquinolone derivatives. When screened for their antimalarial activity by high throughput screening against asexual blood-stage parasites, almost all compounds showed potent and selective sub-micromolar activities. Computational investigation was performed to identify the antiplasmodial potential targets. Ligand-based similarity search predicted the tested compounds to act as hemozoin inhibitors. Computational target identification results were further validated by competitive hemozoin inhibitory properties of hydroxyquinoline and hydroxyquinolone derivatives in vitro. The overall results suggest this natural scaffold is of potential to be developed as antimalarial drug lead.

Metabolomic profiling, biological evaluation of Aspergillus awamori, the river Nile-derived fungus using epigenetic and OSMAC approaches.

LC-HRMS-based metabolomics approach was applied to the river Nile-derived fungus Aspergillus awamori after its fermentation on four different media and using four epigenetic modifiers as elicitors. Thereafter, a comprehensive multivariate statistical analysis such as PCA, PLS-DA and OPLS-DA were employed to explain the generated metabolomic data (1587 features). PCA showed that the fungus displayed a unique chemical profile in each medium or elicitor. Additionally, PLS-DA results revealed the upregulated metabolites under each of these conditions. Results indicated that both rice and malt dextrose agar were recognized as the best media in terms of secondary metabolites diversity and showed better profiles than the four applied epigenetic modifiers, of which nicotinamide was the best secondary metabolite elicitor. Testing the antibacterial and cytotoxic effects of all A. awamori-derived extracts revealed that using epigenetic modifiers can induce antimicrobial metabolites against S. aureus and E. coli, whereas using rice, malt dextrose or nicotinamide can induce groups of cytotoxic metabolites. OPLS-DA results assisted in the putative identification of the induced metabolites that could be responsible for these observed inhibitory activities. This study highlighted how powerful the OSMAC approach in maximizing of the chemical diversity of a single organism. Furthermore, it revealed the power of metabolomics in tracing, profiling and categorizing such chemical diversity and even targeting the possible bioactive candidates which require further scaling up studies in the future.

Application of metabolomics and molecular networking in investigating the chemical profile and antitrypanosomal activity of British bluebells (Hyacinthoides non-scripta).

Bulb, leaf, scape and flower samples of British bluebells (Hyacinthoides non-scripta) were collected regularly for one growth period. Methanolic extracts of freeze-dried and ground samples showed antitrypanosomal activity, giving more than 50% inhibition, for 20 out of 41 samples. High-resolution mass spectrometry was used in the dereplication of the methanolic extracts of the different plant parts. The results revealed differences in the chemical profile with bulb samples being distinctly different from all aerial parts. High molecular weight metabolites were more abundant in the flowers, shoots and leaves compared to smaller molecular weight ones in the bulbs. The anti-trypanosomal activity of the extracts was linked to the accumulation of high molecular weight compounds, which were matched with saponin glycosides, while triterpenoids and steroids occurred in the inactive extracts. Dereplication studies were employed to identify the significant metabolites via chemotaxonomic filtration and considering their previously reported bioactivities. Molecular networking was implemented to look for similarities in fragmentation patterns between the isolated saponin glycoside at m/z 1445.64 [M + formic-H]- equivalent to C64H104O33 and the putatively found active metabolite at m/z 1283.58 [M + formic-H]- corresponding to scillanoside L-1. A combination of metabolomics and bioactivity-guided approaches resulted in the isolation of a norlanostane-type saponin glycoside with antitrypanosomal activity of 98.9% inhibition at 20 µM.

A New Bioactive Compound From the Marine Sponge-Derived Streptomyces sp. SBT348 Inhibits Staphylococcal Growth and Biofilm Formation.

Staphylococcus epidermidis, the common inhabitant of human skin and mucosal surfaces has emerged as an important pathogen in patients carrying surgical implants and medical devices. Entering the body via surgical sites and colonizing the medical devices through formation of multi-layered biofilms leads to refractory and persistent device-related infections (DRIs). Staphylococci organized in biofilms are more tolerant to antibiotics and immune responses, and thus are difficult-to-treat. The consequent morbidity and mortality, and economic losses in health care systems has strongly necessitated the need for development of new anti-bacterial and anti-biofilm-based therapeutics. In this study, we describe the biological activity of a marine sponge-derived Streptomyces sp. SBT348 extract in restraining staphylococcal growth and biofilm formation on polystyrene, glass, medically relevant titan metal, and silicone surfaces. A bioassay-guided fractionation was performed to isolate the active compound (SKC3) from the crude SBT348 extract. Our results demonstrated that SKC3 effectively inhibits the growth (MIC: 31.25 µg/ml) and biofilm formation (sub-MIC range: 1.95-<31.25 µg/ml) of S. epidermidis RP62A in vitro. Chemical characterization of SKC3 by heat and enzyme treatments, and mass spectrometry (HRMS) revealed its heat-stable and non-proteinaceous nature, and high molecular weight (1258.3 Da). Cytotoxicity profiling of SKC3 in vitro on mouse fibroblast (NIH/3T3) and macrophage (J774.1) cell lines, and in vivo on the greater wax moth larvae Galleria mellonella revealed its non-toxic nature at the effective dose. Transcriptome analysis of SKC3 treated S. epidermidis RP62A has further unmasked its negative effect on central metabolism such as carbon flux as well as, amino acid, lipid, and energy metabolism. Taken together, these findings suggest a potential of SKC3 as a putative drug to prevent staphylococcal DRIs.

Genomic insights into specialized metabolism in the marine actinomycete Salinispora.

Comparative genomics is providing new opportunities to address the diversity and distributions of genes encoding the biosynthesis of specialized metabolites. An analysis of 119 genome sequences representing three closely related species of the marine actinomycete genus Salinispora reveals extraordinary biosynthetic diversity in the form of 176 distinct biosynthetic gene clusters (BGCs) of which only 24 have been linked to their products. Remarkably, more than half of the BGCs were observed in only one or two strains, suggesting they were acquired relatively recently in the evolutionary history of the genus. These acquired gene clusters are concentrated in specific genomic islands, which represent hot spots for BGC acquisition. While most BGCs are stable in terms of their chromosomal position, others migrated to different locations or were exchanged with unrelated gene clusters suggesting a plug and play type model of evolution that provides a mechanism to test the relative fitness effects of specialized metabolites. Transcriptome analyses were used to address the relationships between BGC abundance, chromosomal position and product discovery. The results indicate that recently acquired BGCs can be functional and that complex evolutionary processes shape the micro-diversity of specialized metabolism observed in closely related environmental bacteria.

Marine Sponge-Derived Streptomyces sp. SBT343 Extract Inhibits Staphylococcal Biofilm Formation.

Staphylococcus epidermidis and Staphylococcus aureus are opportunistic pathogens that cause nosocomial and chronic biofilm-associated infections. Indwelling medical devices and contact lenses are ideal ecological niches for formation of staphylococcal biofilms. Bacteria within biofilms are known to display reduced susceptibilities to antimicrobials and are protected from the host immune system. High rates of acquired antibiotic resistances in staphylococci and other biofilm-forming bacteria further hamper treatment options and highlight the need for new anti-biofilm strategies. Here, we aimed to evaluate the potential of marine sponge-derived actinomycetes in inhibiting biofilm formation of several strains of S. epidermidis, S. aureus, and Pseudomonas aeruginosa. Results from in vitro biofilm-formation assays, as well as scanning electron and confocal microscopy, revealed that an organic extract derived from the marine sponge-associated bacterium Streptomyces sp. SBT343 significantly inhibited staphylococcal biofilm formation on polystyrene, glass and contact lens surfaces, without affecting bacterial growth. The extract also displayed similar antagonistic effects towards the biofilm formation of other S. epidermidis and S. aureus strains tested but had no inhibitory effects towards Pseudomonas biofilms. Interestingly the extract, at lower effective concentrations, did not exhibit cytotoxic effects on mouse fibroblast, macrophage and human corneal epithelial cell lines. Chemical analysis by High Resolution Fourier Transform Mass Spectrometry (HRMS) of the Streptomyces sp. SBT343 extract proportion revealed its chemical richness and complexity. Preliminary physico-chemical characterization of the extract highlighted the heat-stable and non-proteinaceous nature of the active component(s). The combined data suggest that the Streptomyces sp. SBT343 extract selectively inhibits staphylococcal biofilm formation without interfering with bacterial cell viability. Due to absence of cell toxicity, the extract might represent a good starting material to develop a future remedy to block staphylococcal biofilm formation on contact lenses and thereby to prevent intractable contact lens-mediated ocular infections.

An Enrichment of CRISPR and Other Defense-Related Features in Marine Sponge-Associated Microbial Metagenomes.

Many marine sponges are populated by dense and taxonomically diverse microbial consortia. We employed a metagenomics approach to unravel the differences in the functional gene repertoire among three Mediterranean sponge species, Petrosia ficiformis, Sarcotragus foetidus, Aplysina aerophoba and seawater. Different signatures were observed between sponge and seawater metagenomes with regard to microbial community composition, GC content, and estimated bacterial genome size. Our analysis showed further a pronounced repertoire for defense systems in sponge metagenomes. Specifically, clustered regularly interspaced short palindromic repeats, restriction modification, DNA phosphorothioation and phage growth limitation systems were enriched in sponge metagenomes. These data suggest that defense is an important functional trait for an existence within sponges that requires mechanisms to defend against foreign DNA from microorganisms and viruses. This study contributes to an understanding of the evolutionary arms race between viruses/phages and bacterial genomes and it sheds light on the bacterial defenses that have evolved in the context of the sponge holobiont.

The pathology of sponge orange band disease affecting the Caribbean barrel sponge Xestospongia muta.

The aim of this study was to examine sponge orange band (SOB) disease affecting the prominent Caribbean sponge Xestospongia muta. Scanning and transmission electron microscopy revealed that SOB is accompanied by the massive destruction of the pinacoderm. Chlorophyll a content and the main secondary metabolites, tetrahydrofurans, characteristic of X. muta, were significantly lower in bleached than in healthy tissues. Denaturing gradient gel electrophoresis using cyanobacteria-specific 16S rRNA gene primers revealed a distinct shift from the Synechococcus/Prochlorococcus clade of sponge symbionts towards several clades of unspecific cyanobacteria, including lineages associated with coral disease (i.e. Leptolyngbya sp.). Underwater infection experiments were conducted by transplanting bleached cores into healthy individuals, but revealed no signs of SOB development. This study provided no evidence for the involvement of a specific microbial pathogen as an etiologic agent of disease; hence, the cause of SOB disease in X. muta remains unidentified.