Abstract
Sponges, the most primitive multicellular animals, contain a large number of unique microbial communities. Sponge-associated microorganisms, particularly actinomyces, have the potential to produce diverse active natural products. However, a large number of silent secondary metabolic gene clusters have failed to be revived under laboratory culture conditions. In this study, iterative atmospheric room-temperature plasma. (ARTP) mutagenesis coupled with multi-omics conjoint analysis was adopted to activate the inactive wild Streptomyces strain. The desirable exposure time employed in this study was 75 s to obtain the appropriate lethality rate (94%) and mutation positive rate (40.94%). After three iterations of ARTP mutagenesis, the proportion of mutants exhibiting antibacterial activities significantly increased by 75%. Transcriptome analysis further demonstrated that the differential gene expression levels of encoding type I lasso peptide aborycin had a significant upward trend in active mutants compared with wild-type strains, which was confirmed by LC-MS results with a relative molecular mass of 1082.43 ([M + 2H]2+ at m/z = 2164.86). Moreover, metabolome comparative analysis of the mutant and wild-type strains showed that four spectra or mass peaks presented obvious differences in terms of the total ion count or extracting ion current profiles with each peak corresponding to a specific compound exhibiting moderate antibacterial activity against Gram-positive indicators. Taken together, our data suggest that the ARTP treatment method coupled with multi-omics profiling analysis could be used to estimate the valid active molecules of metabolites from microbial crudes without requiring a time-consuming isolation process.
Highlights
Marine sponges are among the oldest Metazoans on Earth (Webster and Taylor, 2012)
The first atmospheric room-temperature plasma. (ARTP) study was a report on Streptomyces avermitilis mutants that produced >40% more avermectins B1a than the wild-type strain (Wang et al, 2010)
More than 40 different microbial species involving 300 ARTP case studies have applied the ARTP mutagenesis method to improve the yield of diverse antibiotics, enzymes, and intermediate active metabolic compounds, in actinomycetes (Ottenheim et al, 2018)
Summary
Marine sponges (phylum Porifera) are among the oldest Metazoans on Earth (Webster and Taylor, 2012). By combining genome mining approaches with antiSMASH tools, bacterial PKs and non-ribosomal peptides (NRPs), as well as complex secondary metabolites with antimicrobial, antiviral, anti-infective, or anticancer properties, have been identified (Lee et al, 2020a) These new characterization methods have predicted new biosynthetic pathways and their related natural compounds. Based on the gene mining, transcriptome, and metabolomic results, the transcriptional expression levels and corresponding metabolites of the BGCs encoding antimicrobial products were comparatively analyzed between mutants and the wild-type strain This may provide valuable guidance for further studies on activating the silent or cryptic gene clusters of actinobacteria, while supporting further research on the related mechanisms of compound biosynthesis and the targeted development of bioactive products
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