Time-Dependent Production of the Bioactive Peptides Endolides A and B and the Polyketide Mariline A from the Sponge-Derived Fungus Stachylidium bicolor 293K04

Abstract

Previous investigations of the sponge-derived fungus Stachylidium bicolor (S. bicolor)293K04 led to the isolation of the biosynthetically unusual polyketides marilines A-C and the bioactive tetrapeptides endolides A-B, identified as potential neuropathic drug leads. Furthermore, prior extended solid cultivation of S. bicolor 293K04 for 60 days resulted in a significant increase of polyketide yield, and the isolation of seven new polyketides. Due to the interest in endolide activity, unusual biosynthetic diversity, and the late stage polyketide production, we studied the cultivation conditions for determining the production time distribution and yields of these secondary metabolites. Results indicated a first production phase of secondary metabolite dominated by peptides, after 21-23 days. Polyketide mariline A1/A2 only started at day 35 of growth, an unusually late period for secondary metabolite expression. This unusual bimodal sequential expression of different families of secondary metabolites suggests value in exploring extended cultivation times to identify novel bioactive fungal compounds.

Unveiling Concealed Functions of Endosymbiotic Bacteria Harbored in the Ascomycete Stachylidium bicolor

“…2) in solid and liquid media following cultivation periods of ca. 40 days (17,25). Here, we isolated from the culture extracts of S. bicolor also the new natural product onychocin D (see details in Fig.…”

“…Culture media were BMS (biomalt with added artificial seawater: Villa Natura Kraftnahr biomalt, 20 g/liter; sea salts [Sigma], 40 g/liter), TSB (Trypticase soy broth; BD, Difco), or oatmeal (BD, Difco). Fungal cultures on liquid media (10 ml, EPA vials) were performed as previously described (25). Conidia from 25-day-old cultures grown on solid oatmeal were gently harvested with a loop using a saline (0.8% NaCl) solution containing 0.0025% Tween 80.…”

“…Organic extracts of microbial cultures. Fungal liquid cultures were extracted as previously described (25). Bacterial liquid cultures (10 ml) were extracted with methyl ethyl ketone (MEK) (2:1) under vigorous shaking; dimethyl sulfoxide (DMSO) was added to the organic fraction (500 l), and MEK was evaporated under N 2 flow; then, the cultures were concentrated 20ϫ from the initial culture volume in DMSO; finally, the supernatants were cleared of cell debris by centrifugation (10 min, 1,467 ϫ g).…”

Antibacterial Effect and Mode of Action of Secondary Metabolites from Fungal Endophyte Associated with Aloe ferox Mill

“…After 7 days, the zones were constant on E. coli (ATCC 25922) and there was insignificant decrease at day 8 and 9 on S. aureus (ATCC 25925). The decrease might be due to autolysis that might have occurred as the nutrients depleted [43]. In addition, at day 7, the inhibitive intensity of the secondary metabolites in the broth implied that the fungus produced active antibacterial metabolites optimally at its stationary phase.…”

A MassQL-Integrated Molecular Networking Approach for the Discovery and Substructure Annotation of Bioactive Cyclic Peptides

“…To further filter for endolide analogs that are supposedly produced in sufficient amounts for isolation and structure elucidation, only nodes with an intensity (sum of precursor ion) greater than 1.5 × 10 6 were retained, as it was shown before that the peak areas of [M + H] + adducts and concentrations of endolides correlate well. 32 Previous yields of 24.6 mg/L after reisolation of the known endolide A (Table S3, Figure S3) indicated that nodes below this threshold would have yields below 0.1 mg/10L. In this way, we generated a MassQL-annotated molecular network with substructural information for all endolides.…”