Using custom-built primers and nanopore sequencing to evaluate CO-utilizer bacterial and archaeal populations linked to bioH2 production.

The microbial community composition of five distinct thermophilic hot springs was effectively described in this work, using broad-coverage nanopore sequencing (ONT MinION sequencer). By examining environmental samples from the same source, but from locations with different temperatures, bioinformatic analysis revealed dramatic changes in microbial diversity and archaeal abundance. More specifically, no archaeal presence was reported with universal bacterial primers, whereas a significant archaea presence and also a wider variety of bacterial species were reported. These results revealed the significance of primer preference for microbiomes in extreme environments. Bioinformatic analysis was performed by aligning the reads to 16S microbial databases for identification using three different alignment methods, Epi2Me (Fastq 16S workflow), Kraken, and an in-house BLAST tool, including comparison at the genus and species levels. As a result, this approach to data analysis had a significant impact on the genera identified, and thus, it is recommended that use of multiple analysis tools to support findings on taxonomic identification using the 16S region until more precise bioinformatics tools become available. This study presents the first compilation of the ONT-based inventory of the hydrogen producers in the designated hot springs in Türkiye.

Comparing the significance of the utilization of next generation and third generation sequencing technologies in microbial metagenomics.

Since the exploration of sequencing began in 2005, third and next-generation sequencing (TGS and NGS) technologies have fundamentally changed metagenomics research. These platforms provide essential benefits regarding speed, cost, quality and precision in the never-ending search for microorganisms' genetic material, regardless of location on earth. TGS are typically represented by technologies driven from power generation by semiconductor chips and utilization of enzymatic reactions by SOLiD/Ion Torrent PGM™ from Life Sciences, sequencing by synthesis using fluorescent labels on HiSeq/MiSeq™ from Illumina, pyrosequencing by GS FLX Titanium/GS Junior from Roche and nanopore-based sequencing by MinION™/GridION™/PromethION™ from Oxford Nanopore Technologies. The evolution of this technology enabled researchers to continually broaden their knowledge of the microbial world. This review presents a comprehensive overview of the recent literature on the utilization of both TGS and NGS technologies for the investigation of microbial metagenomics, their benefits and limitations with real-time examples of novel applications in clinical microbiology and public health, food and agriculture, energy and environment, arts and space.

Supercritical carbon dioxide dried double layer laponite XLS and alginate/polyacrylamide construct and immune response.

Fabrication of immunocompatible tissue constructs for bone-cartilage defect regeneration is of prime importance. In this study, a double layer hydrogel was successfully synthesized, where alginate/polyacrylamide were formulated to represent cartilage layer (5-10 % (w/w) total polymer ratio) and laponite XLS (2-5-8% (w/w))/alginate/polyacrylamide formed bone layer. Hydrogels were dried by supercritical CO2 at 100 and 200 bar, 45 °C, 5 g/min CO2 flow rate for 2 h. Constructs were treated with collagen, then cellularized and embedded in cell-laden GelMA to mimic the cellular microenvironment. The optimum weight ratio of alginate/polyacrylamide:laponite XLS was 10:5 based on mechanical strength test results. The constructs yielded high porosity (91.50 m2/g) and mesoporous structure, owing to the diffusivity of CO2 at 200 bar (0.49 × 10-7 m2/s). Constructs were then treated with collagen to increase cell adhesion and ATDC5 cells were seeded in the cartilage layer, whereas hFOB cells to the bone layer. About 10-15 % higher cell viability was attained. The porous structure of the construct allowed infiltration of macrophages, promoted polarization and positively affected the behavior of macrophages, yielding a decrease in M1 markers, whereas an increase in M2 on day 4. The formulated tissue constructs would be of value in tissue engineering applications.