Riboflavin, an intermediate of primary metabolism, is one kind of important food additive with high economic value. The microbial cell factory Bacillus subtilis has already been proven to possess significant importance for the food industry and have become one of the most widely used riboflavin-producing strains. In the practical fermentation processes, a sharp decrease in riboflavin production is encountered along with a decrease in the dissolved oxygen (DO) tension. Influence of this oxygen availability on riboflavin biosynthesis through carbon central metabolic pathways in B. subtilis is unknown so far. Therefore the unveiled effective metabolic pathways were still an unaccomplished task till present research work. In this paper, the microscopic regulation mechanisms of B. subtilis grown under different dissolved oxygen tensions were studied by integrating 13C metabolic flux analysis, metabolomics and transcriptomics. It was revealed that the glucose metabolic flux through pentose phosphate (PP) pathway was lower as being confirmed by smaller pool sizes of metabolites in PP pathway and lower expression amount of ykgB at transcriptional level. The latter encodes 6-phosphogluconolactonase (6-PGL) under low DO tension. In response to low DO tension in broth, the glucose metabolic flux through Embden–Meyerhof–Parnas (EMP) pathway was higher and the gene, alsS, encoding for acetolactate synthase was significantly activated that may result due to lower ATP concentration and higher NADH/NAD+ ratio. Moreover, ResE, a membrane-anchored protein that is capable of oxygen regulated phosphorylase activity, and ResD, a regulatory protein that can be phosphorylated and dephosphorylated by ResE, were considered as DO tension sensor and transcriptional regulator respectively. This study shows that integration of transcriptomics, 13C metabolic flux analysis and metabolomics analysis provides a comprehensive understanding of biosynthesized riboflavin’s regulatory mechanisms in B. subtilis grown under different dissolved oxygen tension conditions. The two-component system, ResD–ResE, was considered as the signal receiver of DO tension and gene regulator that led to differences between biomass and riboflavin production after triggering the shifts in gene expression, metabolic flux distributions and metabolite pool sizes.

An enclosed air-lift photobioreactor (ALPBR) is considered an efficient lab-scale bioreactor for microalgae cell growth and lipid biosynthesis. However, fluid-induced shear stress and mixing are two main factors that affect physiological metabolism in microalgal cell cultures. Herein, a 50-L ALPBR after being designed and manufactured was evaluated for microalgal suspension culture. Moreover, computational fluid dynamics (CFD) simulation was used to characterize the hydrodynamics of ALPBR. Specifically, two model microalgae species Chlorella vulgaris and Chlorella protothecoides were cultured with aeration rates at 0.1, 0.2, 0.3, 0.4 volume per volume per minute (vvm). The hydrodynamics of ALPBR while considering shear rate, volume average velocity, turbulent energy dissipation rate (EDR), air volume fraction, and mean air bubble diameter, etc. showed that aeration generated a very low shear stress. When C. protothecoides and C. vulgaris were cultured with various aeration rates, the maximum dry cell weights (DCW) of two microalgal species tested were 0.3 vvm and 0.2 vvm, while the maximum lipid contents of both species were about 15.0% of the DCW, which were achieved at an air flow rate of 0.2 vvm. Therefore, we concluded that the shear rate generated by aeration could play a vital role in the cell growth of microalgae grown in an air-lift photobioreactor.

Adipose-derived mesenchymal stem cells (ADMSCs) are considered an efficient and important candidate for male infertility treatment because they contain pluripotent stem cells, which can differentiate into all cells from three germ layers. However, the efficient generation of male germ-like cell (MGLCs) is one of the key issues, and little is known about the mechanisms underlying generation of MGLCs. Herein, we attempt to improve the efficient generation of MGLCs derived during co-culturing of rat ADMSCs with SCs under retinoic acid (RA) and testosterone (T) treatment. ADMSCs isolated from male SD rat were induced into generation of MGLCs by using respective methods in vitro. Transwell insert system was used for co-culturing. Busulfan-induced non-obstructive azoospermia rat mode was used to evaluate spermatogenic recovery ability of treated ADMSCs. Besides, the relative gene expression level was detected by reverse transcription PCR, quantitative RT-PCR. The relative protein expression level was detected by western blot (WB) and immunostaining analysis. The results showed that ADMSCs co-cultured with TM4 cells under RA and T induction enhanced the formation of bigger and tightly packed MGLCs feature colonies in vitro. Moreover, the expression of male germ cell-related markers (Oct4, Stella, Ddx4, Dazl, PGP9.5, Stra8, and ITGα6) is significantly upregulated in TM4 cell-co-cultured ADMSCs in vitro and in busulfan-treated rat testis after injecting TM4 cell-treated ADMSCs for 2 months. Comparatively, the ADMSCs treated by TM4 cell with RA and T exhibited the highest expression of male germ cell-related markers. RA- and T-treated TM4 cell showed fewer dead cells and higher cytokine secretion than untreated groups. The protein expression level of TGFβ-SMAD2/3, JAK2-STAT3, and AKT pathways in ADMSCs co-cultured with TM4 cells under RA and T was higher than others. Whereas, downregulation of male germ cell-related marker expression subsequently inhibited the phosphorylation of SMAD2/3, JAK2, STAT3, and AKT. These results suggested that TM4 cells could efficiently stimulate in vitro generation of MGLCs during co-culturing of ADMSCs under RA and T treatment. Conclusively, the ADMSCs co-cultured with TM4 cell under RA and T induction stimulate the efficient generation of MGLCs in vitro through activating TGFβ-SMAD2/3, JAK2-STAT3, and AKT pathways. Among them, JAK2-STAT3 and AKT pathways are being first reported to show involvement of in vitro generation of MGLCs during ADMSC co-culturing with SCs.

Microbial cell factories (bacteria and fungi) are the leading producers of beneficial natural products such as lycopene, carotene, herbal medicine, and biodiesel etc. These microorganisms are considered efficient due to their effective bioprocessing strategy (monoculture- and consortial-based approach) under distinct processing conditions. Meanwhile, the advancement in genetic and process optimization techniques leads to enhanced biosynthesis of natural products that are known functional ingredients with numerous applications in the food, cosmetic and medical industries. Natural consortia and monoculture thrive in nature in a small proportion, such as wastewater, food products, and soils. In similitude to natural consortia, it is possible to engineer artificial microbial consortia and program their behaviours via synthetic biology tools. Therefore, this review summarizes the optimization of genetic and physicochemical parameters of the microbial system for improved production of natural products. Also, this review presents a brief history of natural consortium and describes the functional properties of monocultures. This review focuses on synthetic biology tools that enable new approaches to design synthetic consortia; and highlights the syntropic interactions that determine the performance and stability of synthetic consortia. In particular, the effect of processing conditions and advanced genetic techniques to improve the productibility of both monoculture and consortial based systems have been greatly emphasized. In this context, possible strategies are also discussed to give an insight into microbial engineering for improved production of natural products in the future. In summary, it is concluded that the coupling of genomic modifications with optimum physicochemical factors would be promising for producing a robust microbial cell factory that shall contribute to the increased production of natural products.