AbstractIn the present study, we relied to a significant extent on the filterability of the smallest microbial cells often referred as ultra-microcells. We well-known surface water microbiomes are unique communities, but their metagenomes have not been extensively mined for new biosynthesis pathways, we don’t even know whether bacterial cells passage through a 0.
1 µm filter membrane are able to produce secondary metabolites. For the first time in this study, we investigated the presence of secondary metabolites in bacterial cells passage through a 0.1 µm filter membrane. Our results demonstrated that these bacterial cells have considerable potential to produce secondary metabolites. We computationally by using antiSMASH and through the use of metagenomics approach identified and analyzed secondary metabolite in an ultra-small bacterial cell. Additionally, in these ultra-small cells, we screened biosynthetic gene clusters with antimicrobial activity including terpene, bacteriocins, non-ribosomal peptide synthases, polyketide synthases. Besides, we performed classification and annotation of KS and C domains, their diversity and phylogeny by NaPDoS.
1. IntroductionBacterial natural systems are varied in different size and some have very small genomes. The existence and important roles of these ultramicrobacteria in microbial communities and ecosystems are confirmed in the various literature 1, 2.
However, a very large gap exists between the ultramicrobacteria, throughput of sequencing and the rate of discovery of novel pathways involved in secondary metabolism predominantly because of these ultra-small bacterial cells are ambiguous, don’t yet fully understand what they do and lack of research work 1, 3. Bacterial secondary metabolites have many different physiochemical, biological properties and have an important role in the development of novel medicines, antitumor drugs and immunosuppressive, antiparasitics drugs. They are the main source for new therapeutic agents and over the 30 years an huge number of drugs on the market are derived from natural products 4, 5. Most antibiotics, such as penicillin, erythromycin or tetracycline, and also other drugs like acarbose(anti-diabetic), artemisinin(anti-malarial), tacrolimus or cyclosporins are derived from natural products 6. For novel compound discovery, microbiologists using sequencing genomes of a wide variety of microbes (the study of a microbial community without the need of cultivation in the laboratory) that describes biodiversity and biochemical potential of a microbial community and full sequence information. There is a limitation to identify the microorganism in cultured isolates 7. The metagenomics approaches have been used to solve this limitation.
Mining of genomes has become a potent, scalable, and cost-effective accessible tool for the natural product research, biosynthesis and drug discovery 8. However, only a few studies were investigated metagenomes databases for biosynthesis of natural product and much more work is needed in this area.The diversity of microbial populations and microbial communities in subsurface environments including water systems exhibit extraordinary phylogenetic diversity and metabolic complexity that has only recently become apparent using culture-independent sequencing-based analytics 9, 10.
Different tools are available for genome mining and also several computational tools have been developed for detection of secondary metabolite biosynthetic pathways in sequenced fungal or bacterial genomes and some of these tools are able to predict substrate specificity of polymerization enzymes, generate approximations for products’ 3D structures, and so on 11.In this paper, we used antiSMASH for identification and analysis genomic of biosynthetic gene clusters. We used antiSMASH, version 4.0, which has undergone numerous improvements 12. And it allows us to run other analyses such as substrate specificity, homologous sequence, predicted structure and more detail information in addition to gene cluster identificationIn the current study we analysis metagenomics of surface water that passed through a 0.
1 ?m membrane filter to extract DNA, metagenome sequencing, assembly, and then using the antiSMASH and NaPDoS. In overall this study aims to (i) mining of metagenomes in ultra-small bacteria, sequence generation (ii) to proof present of secondary metabolite in ultra-small bacterial cell that passes through a 0.1 ?m membrane filter (iii) to screen biosynthesis gene cluster for new complete multi-modular enzymes from PKS and NRPS families, domain classification exploring their diversity and phylogeny.