Abstract
The study of spatial distribution of secondary metabolites within microbial cells facilitates the screening of candidate strains from marine environments for functional metabolites and allows for the subsequent assessment of the production of metabolites, such as antibiotics. This paper demonstrates the first application of Raman microspectroscopy for in situ detection of the antifungal antibiotic amphotericin B (AmB) produced by actinomycetes—Streptomyces nodosus. Raman spectra measured from hyphae of S. nodosus show the specific Raman bands, caused by resonance enhancement, corresponding to the polyene chain of AmB. In addition, Raman microspectroscopy enabled us to monitor the time-dependent change of AmB production corresponding to the growth of mycelia. The Raman images of S. nodosus reveal the heterogeneous distribution of AmB within the mycelia and individual hyphae. Moreover, the molecular association state of AmB in the mycelia was directly identified by observed Raman spectral shifts. These findings suggest that Raman microspectroscopy could be used for in situ monitoring of antibiotic production directly in marine microorganisms with a method that is non-destructive and does not require labeling.
Highlights
Various secondary metabolites, which have broad functions—antibacterial, antifungal, antiviral, antitumor, and antiprotozoal—have been isolated from different microbes found in terrestrial soils and marine sediments [1,2,3]
This study demonstrates the capability of Raman imaging for non-destructive screening of antibiotic producers and the potential of Raman microspectroscopy, as an in-line monitoring technique, to be a tool for use in antibiotic production from industrial scale fermentation cultures
S. nodosus mycelia cultivated under amphotericin B (AmB)-inducing and -non-inducing conditions were placed on an agar plate with
Summary
Various secondary metabolites, which have broad functions—antibacterial, antifungal, antiviral, antitumor, and antiprotozoal—have been isolated from different microbes found in terrestrial soils and marine sediments [1,2,3]. The purified microbial extracts are subsequently analyzed by GC-MS or NMR to determine their chemical formula and abundance These processes are invasive, time-consuming, laborious, and require a substantial amount of microbe cultures. In conjunction with the demand for novel drugs from environmental sources and cost-effective production, a non-destructive technique for compositional analysis of microbial secondary metabolites is required. Since secondary metabolites such as antibiotics generally have diverse and distinctive chemical structures, we postulated that Raman imaging has the potential to distinguish antibiotics from other biomolecules in living cells without labeling or extraction. This study demonstrates the capability of Raman imaging for non-destructive screening of antibiotic producers and the potential of Raman microspectroscopy, as an in-line monitoring technique, to be a tool for use in antibiotic production from industrial scale fermentation cultures
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