Recent advances in the culture-independent discovery of natural products using metagenomic approaches

Natural products derived from bacterial sources have long been pivotal in the discovery of drug leads.However,the cultivation of only about 1%of bacteria in laboratory settings has left a significant portion of biosynthetic diversity hidden within the genomes of uncultured bacteria.Advances in sequencing technologies now enable the exploration of genetic material from these metagenomes through culture-independent methods.This approach involves extracting genetic sequences from environmental DNA and applying a hybrid methodology that combines functional screening,sequence tag-based homology screening,and bioinformatic-assisted chemical synthesis.Through this process,numerous valuable natural products have been identified and synthesized from previously uncharted metagenomic territories.This paper provides an overview of the recent advancements in the utilization of culture-independent techniques for the discovery of novel biosynthetic gene clusters and bioactive small molecules within metagenomic libraries.

Comparing results of cultured and uncultured biological methods used in biological phosphorus removal

Increasing attention has been paid to phosphate-accumulating organisms （PAOs） for their important role in biological phosphorus removal. In this study, microbial communities of PAOs cultivated under different carbon sources （sewage, glucose, and sodium acetate） were investigated and compared through culture-dependent and culture-independent methods, respectively. The results obtained using denaturing gradient gel electrophoresis （DGGE） of polymerase chain reaction-amplified 16S rDNA fragments revealed that the diversity of bacteria in a sewage-fed reactor （1#） was much higher than in a glucose-fed one （2#） and a sodium acetate-fed one （3#）; there were common PAOs in three reactors fed by different carbon sources. Five strains were separated from three systems by using a phosphaterich medium; they were from common bacteria isolated and three isolates could not be found in DGGE profile at all. Two isolates had good phosphorus removal ability. When the microbial diversity was studied, the molecular biological method was better than the culture-dependent one. When phosphorus removal characteristics were investigated, culture-dependent approach was more effective. Thus a combination of two methods is necessary to have a comprehensive view of PAOs.

Bacterial diversity in Arctic marine sediment determined by culture-dependent and-independent approaches

Bacterial diversity in surface sediment from the Arctic Ocean was investigated by culture-dependent and-independent approaches. Conventional culture-dependent techniques revealed 11 strains based on their distinct morphological characteristics on marine Zobell 2216E agar plates. Phylogenetic analysis showed that these isolates belonged to three major lineages of the Bacteria,γ-proteobacteria, Bacteroidetes and Actinobacteria, and that they included 10 genera. Most isolates were psychrotrophic, and NaCl was not necessary for their growth. Furthermore, they exhibited activity of at least one extracellular hydrolytic enzyme at 4＆#176;C and had various abilities to assimilate carbon sources. A total of 67 phylotypes were detected among 142 clones based on the 16S rRNA library of the total community DNA and grouped into nine major lineages of bacteria. Phylotypes afifliated withγ-,δ-andε-proteobacteria accounted for 36.7%, 21.8%and 16.9%of the total clones, respectively. The rest of the clones belonged to Bacteroidetes,α-proteobacteria, Actinobacteria, Fusobacteria, Nitrospirae and an unclassiifed group.

Study on Endophytic Bacteria Diversity in the Root of Peperomia dindygulensis Miq. by Applying Clone Library Construction Method

[Objectives] To study the endophytic bacteria diversity in the root of Peperomia dindygulensis Miq. [Methods] The root of P.dindygulensis Miq. was taken as materials,culture-independent method was applied to build 16 S rDNA clone library for endophytic bacteria of P. dindygulensis Miq. [Results] In the 16 S rDNA clone library for endophytic bacteria of P. dindygulensis Miq.,95 clones separately belonged to 46 OTUs,Phylogenetic analysis indicated that these cloning sequences separately belonged to phylum Proteobacteria,phylum Bacteroidetes,phylum Actinobacteria,phylum Nitrospirae,and phylum Gemmatimonadetes. In these,phylum Proteobacteria accounted for 70. 53% of total number of clones,so it was the most dominant group in this library. [Conclusions]The endophytic bacteria diversity in the root of P.dindygulensis Miq. is rich and this study is expected to provide scientific basis for protection,development and use of resources of P. dindygulensis Miq.

Marine Metagenome as A Resource for Novel Enzymes

More than 99% of identified prokaryotes, including many from the marine environment, cannot be cultured in the laboratory. This lack of capability restricts our knowledge of microbial genetics and community ecology. Metagenomics, the culture-independent cloning of environmental DNAs that are isolated directly from an environmental sample, has already provided a wealth of information about the uncultured microbial world. It has also facilitated the discovery of novel bio- catalysts by allowing researchers to probe directly into a huge diversity of enzymes within natural microbial communities. Recent advances in these studies have led to a great interest in recruiting microbial enzymes for the development of environmentally-friendly industry. Although the metage- nomics approach has many limitations, it is expected to provide not only scientific insights but also economic benefits, especially in industry. This review highlights the importance of metagenomics in mining microbial lipases, as an example, by using high-throughput techniques. In addition, we dis- cuss challenges in the metagenomics as an important part of bioinformatics analysis in big data.