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- SourceAvailable from: Fernando Reyes[Show abstract] [Hide abstract]
ABSTRACT: Forty four marine actinomycetes of the family Microccocaceae isolated from sponges collected primarily in Florida Keys (USA) were selected from our strain collection to be studied as new sources for the production of bioactive natural products. A 16S rRNA gene based phylogenetic analysis showed that the strains are members of the genera Kocuria and Micrococcus. To assess their biosynthetic potential, the strains were PCR screened for the presence of secondary metabolite genes encoding nonribosomal synthetase (NRPS) and polyketide synthases (PKS). A small extract collection of 528 crude extracts generated from nutritional microfermentation arrays was tested for the production of bioactive secondary metabolites against clinically relevant strains (Bacillus subtilis, methicillin-resistant Staphylococcus aureus (MRSA), Acinetobacter baumannii and Candida albicans). Three independent isolates were shown to produce a new anti-MRSA bioactive compound that was identified as kocurin, a new member of the thiazolyl peptide family of antibiotics emphasizing the role of this family as a prolific resource for novel drugs.
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ABSTRACT: A new thiazolyl peptide, kocurin (1), was isolated from culture broths of a marine-derived Kocuria palustris. Its structural elucidation was accomplished using a combination of spectroscopic and chemical methods, including HRMS, extensive 1D and 2D NMR analysis, MS/MS fragmentation, and chemical degradation and Marfey's analysis of the resulting amino acid residues. The structure herein reported corrects that previously assigned to PM181104 (3). Kocurin displayed activity against methicillin-resistant Staphylococcus aureus (MRSA), with MIC values in the submicromolar range.
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ABSTRACT: Methods for manipulating and fermenting microorganisms in multi-well plates offer unlimited possibilities for high-throughput parallel experimentation. Furthermore, bar-coded data tracking and downstream processing with modern liquid handling equipment reduce handling errors and are able to format microbial products for autosampler-equipped analytical instruments, e.g., HPLCs, mass spectrometers, and plate readers. An integrated system for high-throughput culturing of filamentous fungi replicating strains across many fermentation parameters, called nutritional arrays, was developed. It takes advantage of this equipment while addressing the age-old dilemma of how to manipulate fungal phenotypes to express a more complete spectrum of their secondary metabolites. Growth of any given strain in a well-designed nutritional array increases the chances of detecting a biologically active metabolite while reducing the manpower and materials needed for preparing individual fermentations and extracts. Fungi fermented in nutritional arrays are directly processed in a semi-automated fashion and the extracts prepared for bioassays and analytical chemistry. The necessary equipment, custom tools, and protocols to grow fungi in nutritional arrays are described along with examples of bioactive secondary metabolites discovered using this system.
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