Genomic and Chemical Decryption of the Bacteroidetes Phylum for Its Potential to Biosynthesize Natural Products.

With progress in genome sequencing and data sharing, 1,000s of bacterial genomes are publicly available. Genome mining-using bioinformatics tools in terms of biosynthetic gene cluster (BGC) identification, analysis, and rating-has become a key technology to explore the capabilities for natural product (NP) biosynthesis. Comprehensively, analyzing the genetic potential of the phylum Bacteroidetes revealed as the most talented genus in terms of BGC abundance and diversity.

Novel Glycerophospholipid, Lipo- and -acyl Amino Acids from Bacteroidetes: Isolation, Structure Elucidation and Bioactivity.

The 'core' metabolome of the Bacteroidetes genus was recently discovered to consist of only seven metabolites. A structural relationship in terms of shared lipid moieties among four of them was postulated. Here, structure elucidation and characterization via ultra-high resolution mass spectrometry (UHR-MS) and nuclear magnetic resonance (NMR) spectroscopy of those four lipids (two lipoamino acids (LAAs), two lysophosphatidylethanolamines (LPEs)), as well as several other undescribed LAAs and -acyl amino acids (NAAAs), identified during isolation were carried out.

Combination of high-throughput microfluidics and FACS technologies to leverage the numbers game in natural product discovery.

High-throughput platforms facilitating screening campaigns of environmental samples are needed to discover new products of natural origin counteracting the spreading of antimicrobial resistances constantly threatening human and agricultural health. We applied a combination of droplet microfluidics and fluorescence-activated cell sorting (FACS)-based technologies to access and assess a microbial environmental sample. The cultivation performance of our microfluidics workflow was evaluated in respect to the utilized cultivation media by Illumina amplicon sequencing of a pool of millions of droplets, respectively.

High-Throughput Cultivation for the Selective Isolation of Acidobacteria From Termite Nests.

Microbial communities in the immediate environment of socialized invertebrates can help to suppress pathogens, in part by synthesizing bioactive natural products. Here we characterized the core microbiomes of three termite species (genus ) and their nest material to gain more insight into the diversity of termite-associated bacteria. Sampling a healthy termite colony over time implicated a consolidated and highly stable microbiome, pointing toward the fact that beneficial bacterial phyla play a major role in termite fitness.

Selection of sponge-associated bacteria with high potential for the production of antibacterial compounds.

The potential of sponge-associated bacteria for the biosynthesis of natural products with antibacterial activity was evaluated. In a preliminary screening 108 of 835 axenic isolates showed antibacterial activity. Active isolates were identified by 16S rRNA gene sequencing and selection of the most promising strains was done in a championship like approach, which can be done in every lab and field station without expensive equipment.

Optimizing the composition of a synthetic cellulosome complex for the hydrolysis of softwood pulp: identification of the enzymatic core functions and biochemical complex characterization.

Background: The development of efficient cellulase blends is a key factor for cost-effectively valorizing biomass in a new bio-economy. Today, the enzymatic hydrolysis of plant-derived polysaccharides is mainly accomplished with fungal cellulases, whereas potentially equally effective cellulose-degrading systems from bacteria have not been developed. Particularly, a thermostable multi-enzyme cellulase complex, the cellulosome from the anaerobic cellulolytic bacterium is promising of being applied as cellulolytic nano-machinery for the production of fermentable sugars from cellulosic biomass.

A metagenome-derived thermostable β-glucanase with an unusual module architecture which defines the new glycoside hydrolase family GH148.

The discovery of novel and robust enzymes for the breakdown of plant biomass bears tremendous potential for the development of sustainable production processes in the rapidly evolving new bioeconomy. By functional screening of a metagenomic library from a volcano soil sample a novel thermostable endo-β-glucanase (EngU) which is unusual with regard to its module architecture and cleavage specificity was identified. Various recombinant EngU variants were characterized.

Comparative characterization of all cellulosomal cellulases from reveals high diversity in endoglucanase product formation essential for complex activity.

Background: is a paradigm for efficient cellulose degradation and a promising organism for the production of second generation biofuels. It owes its high degradation rate on cellulosic substrates to the presence of supra-molecular cellulase complexes, cellulosomes, which comprise over 70 different single enzymes assembled on protein-backbone molecules of the scaffold protein CipA.

Results: Although all 24 single-cellulosomal cellulases were described previously, we present the first comparative catalogue of all these enzymes together with a comprehensive analysis under identical experimental conditions, including enzyme activity, binding characteristics, substrate specificity, and product analysis.

Functional Screening of Hydrolytic Activities Reveals an Extremely Thermostable Cellulase from a Deep-Sea Archaeon.

Extreme habitats serve as a source of enzymes that are active under extreme conditions and are candidates for industrial applications. In this work, six large-insert mixed genomic libraries were screened for hydrolase activities in a broad temperature range (8-70°C). Among a variety of hydrolytic activities, one fosmid clone, derived from a library of pooled isolates of hyperthermophilic archaea from deep sea vents, displayed hydrolytic activity on carboxymethyl cellulose substrate plates at 70°C but not at lower temperatures.

Characterization of chromosomal and megaplasmid partitioning loci in Thermus thermophilus HB27.

Background: In low-copy-number plasmids, the partitioning loci (par) act to ensure proper plasmid segregation and copy number maintenance in the daughter cells. In many bacterial species, par gene homologues are encoded on the chromosome, but their function is much less understood. In the two-replicon, polyploid genome of the hyperthermophilic bacterium Thermus thermophilus, both the chromosome and the megaplasmid encode par gene homologues (parABc and parABm, respectively).

Identification of novel esterase-active enzymes from hot environments by use of the host bacterium Thermus thermophilus.

Functional metagenomic screening strategies, which are independent of known sequence information, can lead to the identification of truly novel genes and enzymes. Since E. coli has been used exhaustively for this purpose as a host, it is important to establish alternative expression hosts and to use them for functional metagenomic screening for new enzymes.

Genetic analysis of lipolytic activities in Thermus thermophilus HB27.

The extremely thermophilic bacterium Thermus thermophilus HB27 displays lipolytic activity for the hydrolysis of triglycerides. In this study we performed a mutational in vivo analysis of esterases and lipases that confer growth on tributyrin. We interrupted 10 ORFs suspected to encode lipolytic enzymes.

Toxicity of indoxyl derivative accumulation in bacteria and its use as a new counterselection principle.

In this work we describe the conditional toxic effect of the expression of enzymes that cleave 5-bromo-4-chloro-3-indolyl (BCI) substrates and its use as a new counterselection principle useful for the generation of clean and unmarked mutations in the genomes of bacteria. The application of this principle was demonstrated in the thermophile Thermus thermophilus HB27 and in a mesophile for which currently no counterselection markers are available, Micrococcus luteus ATCC 27141. For T.

Screening and expression of genes from metagenomes.

Microorganisms are the most abundant and widely spread organisms on earth. They colonize a huge variety of natural and anthropogenic environments, including very specialized ecological niches and even extreme habitats, which are made possible by the immense metabolic diversity and genetic adaptability of microbes. As most of the organisms from environmental samples defy cultivation, cultivation-independent metagenomics approaches have been applied since more than one decade to access and characterize the phylogenetic diversity in microbial communities as well as their metabolic potential and ecological functions.