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Background: Inner gene therapy offers great promises as a potential treatment for hearing loss. Aims/objectives: One of the critical determinants of the success of inner ear gene therapy is to find a delivery method which results in consistent transduction efficiency of targeted cell types while minimizing hearing loss. Material and methods: Surgery was performed only in the right ear of each Bama miniature pig, and the left ear served as a control. The gene delivery to inner ear via round window membrane (RWM) and posterior semicircular canal (PSC) approach was performed with the viral vector AAV1-CMV-GFP. Results: The gene delivery through RWM and the PSC (canalostomy) is able to perfuse the inner ear. Conclusions and significance: The easy anatomic identification of the PSC, as to RWM, as well as minimal manipulation of the temporal bone required, make this surgical approach an attractive option for inner ear gene delivery in big animal model. [ABSTRACT FROM AUTHOR]

Highlights • Fermentation kinetics of DHA production using Schizochytrium sp. was developed. • Growth-uncoupled DHA production by Schizochytrium sp. was studied. • Multi-stage continuous fermentation prediction model was established and applied. • The DHA efficient production was realized by multi-stage continuous fermentation. Abstract Docosahexaenoic acid (DHA) has wide-ranging benefits for normal development of the visual and nervous systems in infants. A sustainable source of DHA production through fermentation using Schizochytrium sp. has been developed. In this paper, we present the discovery of growth-uncoupled DHA production by Schizochytrium sp. and the development of corresponding kinetic models of fed-batch fermentations, which can be used to describe and predict the cell growth and substrate utilization as well as lipid and DHA production. Based on this kinetic model, a predictive model of multi-stage continuous fermentation process was established and used to analyze, optimize and design the process parameters. Optimal predicted processes of two-stage and three-stage continuous fermentation were developed and verified in lab-scale bioreactor based on the predicted process parameters. A successful three-stage continuous fermentation was achieved, which increased the lipid, DHA content and DHA productivity by 47.6, 64.3 and 97.1%, respectively, compared with two-stage continuous fermentation. [ABSTRACT FROM AUTHOR]

Lipids rich in polyunsaturated fatty acids are important nutrients. They are traditionally extracted from animals and plants but alternatively can be obtained from microbes through microbial lipid biotechnology. To make this process more economical, apart from strain engineering, the next frontier is through bioprocess and downstream innovation. [ABSTRACT FROM AUTHOR]

Process scale-up is a critical factor for realizing the industrial-scale production of docosahexaenoic acid (DHA). To improve the efficiency of scale-up, a strategy based on computational fluid dynamics analysis and investigation of microbial physiological properties was developed. We first investigated the physiological response of Schizochytrium sp. to different scale bioreactors, and the results indicated that a homogeneous culture environment was beneficial for cell growth and DHA biosynthesis. Fluid dynamics of different scale bioreactors were further investigated and impeller combinations were designed and optimized by computational methods to obtain a homogeneous culture environmental in a 7000 L bioreactor. The results showed that the optimal impeller combination could provide a more balanced culture environment, similar to lab-scale bioreactors. When the optimal strategy was used to scale-up the DHA fermentation, the biomass, DHA concentration, DHA percentage in biomass and DHA productivity reached 128.3 g/L, 39.2 g/L, 30.5%, 326.5 mg/(L h), respectively. This study thus introduces a highly efficient and economical scale-up strategy for DHA production using Schizochytrium sp. [ABSTRACT FROM AUTHOR]

Oxygen uptake rate (OUR) and respiratory quotient (RQ) are key respiratory parameters for docosahexaenoic acid (DHA) production by Schizochytrium sp. HX-308 under dissolved oxygen limited conditions. To investigate the relationship of OUR and RQ with culture status, three independent cultures with different aeration rates were performed in a 50 L bioreactor. OUR was found to be positively correlated with the aeration rate, which reflected the oxygen supply level in each culture. The highest biomass, reaching 124.5 g/L, was achieved under the highest OUR. DHA content was found to be highly correlated with the RQ value, and the highest DHA content (44.85% in total fatty acids, w/w) was achieved in the highest RQ level, which implies that the polyketide synthase pathway was more active. OUR and RQ, which reflect the physiological state of microorganisms, are suggested as synergistic real-time bioprocess monitoring parameters for DHA fermentation. [ABSTRACT FROM AUTHOR]

Polyunsaturated fatty acids (PUFAs) are important for human health. They are traditionally extracted from animals and plants but can be alternatively derived from oleaginous microbes, and engineering microbial metabolism can improve PUFA accumulation. The next frontier is to engineer more efficient PUFA-producing microbes using systems and synthetic biology tools. [ABSTRACT FROM AUTHOR]

Many microorganisms could naturally produce ( R, R)-2,3-butanediol (( R, R)-2,3-BD), which has unique applications due to its special chiral group and spatial configuration. But the low enantio-purity of the product hindered the development of large-scale production. In this work, a synthetic constitutive metabolic pathway for enantiomerically pure ( R, R)-2,3-BD biosynthesis was constructed in Escherichia coli with vector pUC6S, which does not contain any lac sequences. The expression of this artificial constructed gene cluster was optimized by using two different strength of promoters (AlperPLTet01 (P) and AlperBB (P)). The strength of P is twice stronger than P. The fermentation results suggested that the yield of ( R, R)-2,3-BD was higher when using the stronger promoter. Compared with the wild type, the recombinant strain E. coli YJ2 produced a small amount of acetic acid and showed higher glucose consumption rate and higher cell density, which indicated a protection against acetic acid inhibition. In order to further increase the ( R, R)-2,3-BD production by reducing the accumulation of its precursor acetoin, the synthetic operon was reconstructed by adding the strong promoter P in front of the gene ydjL coding for the enzyme of ( R, R)-2,3-BD dehydrogenase which catalyzes the conversion of acetoin to ( R, R)-2,3-BD. The engineered strain E. coli YJ3 showed a 20 % decrease in acetoin production compared with that of E. coli YJ2. After optimization the fermentation conditions, 30.5 g/L of ( R, R)-2,3-BD and 3.2 g/L of acetoin were produced from 80 g/L of glucose within 18 h, with an enantio-purity over 99 %. [ABSTRACT FROM AUTHOR]

Non-conventional yeasts have attracted increasing interest due to their biochemical characteristics and potential applications. Yarrowia lipolytica is a non-conventional yeast with specific characteristics and physiology. The potential physiological and metabolic capabilities of Y. lipolytica , which can assimilate many different carbon sources, including typical hydrophilic and hydrophobic materials, are reviewed in this paper. Concerning the uptake and metabolism substrates, this review focuses particularly on low-cost raw materials, such as glycerol. Moreover, this review presents the results of safety studies of Y. lipolytica . Finally, the wide applications of Y. lipolytica , such as functional enzyme production, metabolite synthesis and environmental bioremediation, are reviewed in this paper. Recently, with the development of system biology and synthetic biology, it was concluded that these technologies will provide new opportunities for potential applications of Y. lipolytica in the future. [ABSTRACT FROM AUTHOR]

Quercetin is a classic flavonoid that inhibits the epithelial–mesenchymal transition (EMT) of tumor cells. However, the effects of quercetin on the EMT of renal tubular epithelial cells, a potential mechanism of renal fibrosis and important characteristic of diabetic nephropathy (DN), remain largely unknown. In the present study, we investigated the effects of quercetin on the EMT of two lines of renal tubular proximal epithelial cells (HK-2 and NRK-52E) induced with high glucose and renal fibrosis resulting from type 1 diabetes and tried to clarify the specific mechanisms underlying these effects. The in vitro results showed that the EMT of HK-2 and NRK-52E cells was induced by high glucose, and mTORC1/p70S6K was highly activated in these two cell lines cultured under high glucose. Quercetin effectively ameliorated the high glucose-induced EMT of HK-2 and NRK-52E cells and inhibited the activation of mTORC1/p70S6K. In vivo, diabetic rats showed a significant decline in renal function and severe renal fibrosis at 14 weeks after STZ injection. Furthermore, mTORC1/p70S6K was activated in the renal cortex of diabetic rats. Treatment with quercetin alleviated the decline in renal function, and the progression of renal fibrosis and inhibited mTORC1/p70S6K activation in the diabetic renal cortex. In addition, we examined the protein and mRNA levels of four transcriptional factors (snail, slug, twist and ZEB-1), which regulate E-cadherin expression at the transcriptional level both in vivo and in vitro. The results revealed that the elevated expression of snail and twist in HK-2 and NRK-52E cells cultured under high glucose and in the renal cortex of diabetic rats was inhibited by quercetin. These results demonstrated that quercetin ameliorates the EMT of HK-2 and NRK-52E cells induced by high glucose and renal fibrosis induced by diabetes, and these effects have been associated with the inhibition of the two transcriptional factors (snail and twist) and the activation of mTORC1/p70S6K. [ABSTRACT FROM AUTHOR]

Nylons are important polyamide (PA) materials that can be polymerized from different monomers. Bio-based nylon monomers are traditionally obtained through chemical conversion from vegetable oils, but they can be more sustainably obtained through multienzymatic catalysis. For large-scale application of this process, enzyme engineering and process innovation must be combined. [ABSTRACT FROM AUTHOR]

Arachidonic acid (ARA)-rich oil production by Mortierella alpina is a long fermentation period needed process due to the low growth rate of the filamentous fungus used. This causes the low productivity of ARA-rich oil and hinders its industrial mass scale production. In the present study, different fed-batch strategies were conducted to shorten the fermentation period. The result showed that compared with the batch culture, the fermentation period was shortened from 7 days to 5 days with the productivity of ARA-rich oil increased from 0.9 g/(L·d) to 1.3 g/(L·d) by using the fed-batch fermentation strategy. Furthermore, repeated fed-batch fermentation strategy was adopted to achieve the purpose of continuous production. By using this strategy, the fermentation period was shortened from 40 days to 26 days in a four cycle repeated fed-batch fermentation. This strategy proved to be convenient and economical for ARA-rich oil commercial production process. [ABSTRACT FROM AUTHOR]

Fungal arachidonic acid (ARA)-rich oil is an important microbial oil that affects diverse physiological processes that impact normal health and chronic disease. In this article, the historic developments and technological achievements in fungal ARA-rich oil production in the past several years are reviewed. The biochemistry of ARA, ARA-rich oil synthesis and the accumulation mechanism are first introduced. Subsequently, the fermentation and downstream technologies are summarized. Furthermore, progress in the industrial production of ARA-rich oil is discussed. Finally, guidelines for future studies of fungal ARA-rich oil production are proposed in light of the current progress, challenges and trends in the field. [ABSTRACT FROM AUTHOR]

Abstract: 2,3-Butanediol is a promising bulk chemical due to its extensive industry applications. The state-of-the-art nature of microbial 2,3-butanediol production is reviewed in this paper. Various strategies for efficient and economical microbial 2,3-butanediol production, including strain improvement, substrate alternation, and process development, are reviewed and compared with regard to their pros and cons. This review also summarizes value added derivatives of biologically produced 2,3-butanediol and different strategies for downstream processing. The future prospects of microbial 2,3-butanediol production are discussed in light of the current progress, challenges, and trends in this field. Guidelines for future studies are also proposed. [Copyright &y& Elsevier]

Microbial preference for glucose implies incomplete and/or slow utilization of lignocellulose hydrolysates, which is caused by the regulatory mechanism named carbon catabolite repression (CCR). In this study, a 2,3-butanediol (2,3-BD) producing Klebsiella oxytoca strain was engineered to eliminate glucose repression of xylose utilization. The crp(in) gene, encoding the mutant cyclic adenosine monophosphate (cAMP) receptor protein CRP(in), which does not require cAMP for functioning, was characterized and overexpressed in K. oxytoca. The engineered recombinant could utilize a mixture of glucose and xylose simultaneously, without CCR. The profiles of sugar consumption and 2,3-BD production by the engineered recombinant, in glucose and xylose mixtures, were examined and showed that glucose and xylose could be consumed simultaneously to produce 2,3-BD. This study offers a metabolic engineering strategy to achieve highly efficient utilization of sugar mixtures derived from the lignocellulosic biomass for the production of bio-based chemicals using enteric bacteria. [ABSTRACT FROM AUTHOR]

Abstract: An industrial medium containing urea as a sole nitrogen source, low levels of corn steep liquor and mineral salts as nutrition factors to retain high 2,3-butanediol production through co-fermentation of glucose and xylose (2:1, wt/wt) by Klebsiella oxytoca was developed. Urea and corn steep liquor were identified as the most significant factors by the two-level Plackett–Burman design. Steepest ascent experiments were applied to approach the optimal region of the two factors and a central composite design was employed to determine their optimal levels. Under the optimal medium, the yield of 2,3-butanediol plus acetoin relative to glucose and xylose was up to 0.428g/g, which was 85.6% of theoretical value. The cheap nitrogen source and nutrition factors combining the co-fermentation process using lignocellulose derived glucose and xylose as the carbon source in the developed medium would be a potential solution to improve the economics of microbial 2,3-butanediol production. [Copyright &y& Elsevier]

Batch fermentative production of 2,3-butanediol by Klebsiella oxytoca was investigated using various oxygen supply methods though varying agitation speed. Based on the analysis of three kinetic parameters including specific cell growth rate (μ), specific glucose consumption rate (qs ) and specific 2,3-butanediol formation rate (qp ), a two-stage agitation speed control strategy, aimed at achieving high concentration, high yield and high productivity of 2,3-butanediol, was proposed. At the first 15h, agitation speed was controlled at 300rpm to obtain high μ for cell growth, subsequently agitation speed was controlled at 200rpm to maintain high qp for high 2,3-butanediol accumulation. Finally, the maximum concentration of 2,3-butanediol reached 95.5gl−1 with the yield of 0.478gg−1 and the productivity of 1.71gl−1 h−1, which were 6.23%, 6.22% and 22.14% over the best results controlled by constant agitation speeds. [Copyright &y& Elsevier]

Xylo-oligosaccharides (XOS) are considered as a promising type of prebiotics that can be used in foods, feeds, and healthcare products. Xylanases play a key role in the production of XOS from xylan. In this study, we conducted a metagenomic analysis of the fecal microbiota from dairy cows fed with different types of fodders. Despite the diversity in their diets, the main phyla observed in all fecal microbiota were Firmicutes and Bacteroidetes. At the genus level, one group of dairy cows that were fed probiotic fermented herbal mixture-containing fodders displayed decreased abundance of Methanobrevibacter and increased growth of beneficial Akkermansia bacteria. Additionally, this group exhibited a high microbial richness and diversity. Through our analysis, we obtained a comprehensive dataset comprising over 280,000 carbohydrate-active enzyme genes. Among these, we identified a total of 163 potential xylanase genes and subsequently expressed 34 of them in Escherichia coli. Out of the 34 expressed genes, two alkaline xylanases with excellent temperature stability and pH tolerance were obtained. Notably, CDW-xyl-8 exhibited xylanase activity of 96.1 ± 7.5 U/mg protein, with an optimal working temperature of 55 ℃ and optimal pH of 8.0. CDW-xyl-16 displayed an activity of 427.3 ± 9.1 U/mg protein with an optimal pH of 8.5 and an optimal temperature at 40 ℃. Bioinformatic analyses and structural modeling suggest that CDW-xyl-8 belongs to GH10 family xylanase, and CDW-xyl-16 is a GH11 family xylanase. Both enzymes have the ability to hydrolyze beechwood xylan and produce XOS. In conclusion, this metagenomic study provides valuable insights into the fecal microbiota composition of dairy cows fed different fodder types, revealing main microbial groups and demonstrating the abundance of xylanases. Furthermore, the characterization of two novel xylanases highlights their potential application in XOS production. [ABSTRACT FROM AUTHOR]

The biocatalytic asymmetric reduction of prochiral ketones for the production of enantiopure alcohols is highly desirable due to its inherent advantages over chemical methods. In this study, a new bacterial strain capable of transforming ketones to corresponding alcohols with high activity and excellent enantioselectivity was discovered in a soil sample. The strain was subsequently identified as Bacillus cereus TQ-2 based on its physiological characteristics and 16S rDNA sequence analysis. Under optimized reaction conditions, the resting cells of B. cereus TQ-2 converted acetophenone to enantioenriched (R)-1-phenylethanol with 99% enantiometric excess following anti-Prelog's rule, which is scarce in biocatalytic ketone reduction. The optimum temperature for the cells was 30 °C, and considerable catalytic activity was observed over a broad pH range from 5.0 to 9.0. The cells showed enhanced catalytic activity in the presence of 15% (v/v) glycerol as a co-substrate. The catalytic activity can also be substantially improved by adding Ca2+ or K+ ions. Moreover, the B. cereus TQ-2 cell was highly active in reducing several structurally diverse ketones and aldehydes to form corresponding alcohols with good to excellent conversion. Our study provides a versatile whole-cell biocatalyst that can be used in the asymmetric reduction of ketones for the production of chiral alcohol, thereby expanding the biocatalytic toolbox for potential practical applications. [ABSTRACT FROM AUTHOR]

Background: α-Humulene is an important biologically active sesquiterpene, whose heterologous production in microorganisms is a promising alternative biotechnological process to plant extraction and chemical synthesis. In addition, the reduction of production expenses is also an extremely critical factor in the sustainable and industrial production of α-humulene. In order to meet the requirements of industrialization, finding renewable substitute feedstocks such as low cost or waste substrates for terpenoids production remains an area of active research. Results: In this study, we investigated the feasibility of peroxisome-engineering strain to utilize waste cooking oil (WCO) for high production of α-humulene while reducing the cost. Subsequently, transcriptome analysis revealed differences in gene expression levels with different carbon sources. The results showed that single or combination regulations of target genes identified by transcriptome were effective to enhance the α-humulene titer. Finally, the engineered strain could produce 5.9 g/L α‐humulene in a 5‐L bioreactor. Conclusion: To the best of our knowledge, this is the first report that converted WCO to α-humulene in peroxisome-engineering strain. These findings provide valuable insights into the high-level production of α-humulene in Y. lipolytica and its utilization in WCO bioconversion. [ABSTRACT FROM AUTHOR]

Oxygen-mediated cell damage is an important issue in aerobic fermentation. In order to counteract these problems, effect of ascorbic acid on cell growth and docosahexaenoic acid (DHA) production was investigated in Schizochytrium sp. Addition of 9 g/L ascorbic acid resulted in 16.16% and 30.44% improvement in cell dry weight (CDW) and DHA yield, respectively. Moreover, the total antioxidant capacity (T-AOC) of cells decreased from 2.17 at 12 h to 0 at 60 h and did not recover, while ascorbic acid addition could extend the time of arrival zero with the reduced intracellular ROS. However, ROS levels still increased after 72 h. Therefore, to further solve the problem of high ROS levels and low T-AOC of cells after 72 h, a two-point addition strategy was proposed. With this strategy, DHA yield was further increased to 38.26 g/L. This work innovatively investigated the feasibility of manipulating Schizochytrium sp. cultivation through ROS level and T-AOC. [ABSTRACT FROM AUTHOR]

In the era of sustainable development, the use of cell factories to produce various compounds by fermentation has attracted extensive attention; however, industrial fermentation requires not only efficient production strains, but also suitable extracellular conditions and medium components, as well as scaling-up. In this regard, the use of biological models has received much attention, and this review will provide guidance for the rapid selection of biological models. This paper first introduces two mechanistic modeling methods, kinetic modeling and constraint-based modeling (CBM), and generalizes their applications in practice. Next, we review data-driven modeling based on machine learning (ML), and highlight the application scope of different learning algorithms. The combined use of ML and CBM for constructing hybrid models is further discussed. At the end, we also discuss the recent strategies for predicting bioreactor scale-up and culture behavior through a combination of biological models and computational fluid dynamics (CFD) models. [ABSTRACT FROM AUTHOR]

[Display omitted] • Comprehensive analysis on the superiority of Y. lipolytica. • Up-to-date overview of synthetic biology tools in Y. lipolytica. • First review on synthesis of diverse natural products by Y. lipolytica. • In-depth discussion on challenges and future perspectives for the production of natural products. With the rapid development of synthetic biology, researchers can design, modify, or even synthesize microorganisms de novo , and microorganisms endowed with unnatural functions can be considered "artificial life" and facilitate the development of functional products. Based on this concept, researchers can solve critical problems related to the insufficient supply of natural products, such as low yields, long production cycles, and cumbersome procedures. Due to its superior performance and unique physiological and biochemical characteristics, Yarrowia lipolytica is a favorable chassis cell used for green biomanufacturing by numerous researchers. This paper mainly reviews the development of synthetic biology techniques for Y. lipolytica and summarizes the recent research progress on the synthesis of natural products in Y. lipolytica. This review will promote the continued innovative development of Y. lipolytica by providing theoretical guidance for research on the biosynthesis of natural products. [ABSTRACT FROM AUTHOR]

Adaptive laboratory evolution (ALE) is an effective method in changing the strain characteristics. Here, ALE with high oxygen as a selection pressure was applied to improve the production capacity of Schizochytrium sp. Results showed that cell dry weight (CDW) of endpoint strain was 32.4% higher than that of starting strain. But slight lipid accumulation impairment was observed. These major performance changes were accompanied with enhanced isocitrate dehydrogenase enzyme activity and reduced ATP:citrate lyase enzyme activity. And a serious decrease of 62.6% in S DHA 140rpm → 170rpm was observed in the endpoint strain. To further study the docosahexaenoic acid (DHA) production ability of evolved strain, fed-batch strategy was applied and 84.34 g/L of cell dry weight and 26.40 g/L of DHA yield were observed. In addition, endpoint strain produced greatly less squalene than starting strain. This work demonstrated that ALE may be a promising tool in modifying microalga strains. [ABSTRACT FROM AUTHOR]

Highlights: [•] Formation and mobilization of lipid fractions were investigated thoroughly. [•] Fatty acid combined on PLs could be changed more easily than NLs. [•] The shift of NL to PL and the increase of UMs happed in lipid turnover stage. [•] DHA enrichment in lipid turnover stage mainly happened in PLs. [•] A positive correlation exited between the synthesis of PUFA and UMs. [ABSTRACT FROM AUTHOR]

The efficient valorization of lignin-derived vanillyl alcohol for the production of value-added biovanillin with enzyme or whole-cell based biotransformation is attracting increasing attention. In this study, a recombinant Escherichia coli HMFO was explored for the production of vanillin through the selective aerobic oxidation of vanillyl alcohol. The resting cells of this strain indicated a high vanillin tolerance and good biocatalytic activity over a temperature range of 10–35 °C. The vanillyl alcohol oxidation process produced vanillin as the sole product, indicating excellent reaction selectivity. Vanillyl alcohol substrate of 250 mM was converted to vanillin under optimal conditions with 96 % yield. In addition, 449.2 mM vanillin was obtained in a fed-batch experiment, providing a productivity of 1.2 g/L per hour and 90 % yield. The production of biovanillin could be further improved by using enzyme engineering and process engineering methods. [Display omitted] • Highly vanillin-tolerant recombinant E.coli HMFO was used for vanillin synthesis. • Vanillin was biosynthesized from lignin-based vanillyl alcohol with E.coli HMFO. • The sole product of vanillin from vanillyl alcohol oxidation was obtained. • Vanillyl alcohol (250 mM) was converted to vanillin with a yield of 96 %. [ABSTRACT FROM AUTHOR]

• D. xibeiensis R13 cells catalyzed the oxidation of aldehydes to carboxylic acids. • Carboxylic acid as a sole product was obtained from the aldehyde oxidation. • D. xibeiensis R13 resting cells demonstrated excellent organic solvent tolerance. • Bioconversion of 250 mM furfural to 2-furoic acid with a yield of 93 % was achieved. The conversion of aldehydes into carboxylic acids through biocatalysis is a promising approach in the fields of green organic synthesis and biotechnology. In this work, we have developed a biocatalytic method for the oxidation of aldehydes to form carboxylic acid in the aqueous phase by using a newly discovered whole D. xibeiensis R13 cell biocatalyst. The whole-cell biocatalyst exhibited excellent organic solvent tolerance and a low biocatalyst/substrate ratio in the selective oxidation of biomass-derived furfural to 2-furoic acid. Under optimal conditions, a yield of 93 % was achieved after 12 h with a conversion of 250 mM furfural substrate. The microbial oxidation of furfural was further scaled up by a fed-batch strategy, with a productivity of 1.3 g/L per hour and a total yield of 83 %. A panel of aldehydes with various reactive moieties was efficiently converted with expected carboxylic acids as the sole product under mild reaction conditions, indicating excellent chemoselectivity (>99 %) and wide substrate scope. The new biocatalytic method avoids the use of toxic transition-metal reagents, representing a green and sustainable alternative to chemical methods. [Display omitted] [ABSTRACT FROM AUTHOR]

The sesquiterpene (−)-α-bisabolol has been shown to have antibacterial, antiseptic, and anti-inflammatory activities, as well as skin-soothing properties, which has led to its wide use in cosmetics. However, the current production of (−)-α-bisabolol via steam-distillation of plants is not sustainable and cannot meet the increasing market demand. In the present study, a sustainable strategy for (−)-α-bisabolol production was developed by constructing a Yarrowia lipolytica cell factory. The oleaginous yeast Y. lipolytica has a native mevalonate pathway and strong acetyl-CoA flux, which can supply adequate precursors for (−)-α-bisabolol production. The endogenous mevalonate pathway and heterologous (−)-α-bisabolol synthesis pathway were optimized by strengthening the rate-limiting enzymes, downregulating competing pathway, as well as balancing the distribution of the common precursor acetyl-CoA between endogenous lipids and heterologous (−)-α-bisabolol synthesis by strengthening the β-oxidation pathway. Finally, (−)-α-bisabolol production of up to 364.23 mg L−1 was achieved in batch culture. This sustainable method has potential for industrial (−)-α-bisabolol production. [ABSTRACT FROM AUTHOR]

Objective: The objectives of this study were to examine the clinical profile of critically ill patients with septic acute kidney injury (AKI) and to investigate clinical characteristics associated with the outcome of patients. Methods: Data from 582 critically ill patients were collected and retrospectively reviewed. Patients were divided into two groups: without AKI development and with AKI development. Baseline characteristics, laboratory, and other clinical data were compared between these two groups, and correlations between the characteristics and AKI development were examined. Patients with AKI development were further divided into two groups according to the survival outcome, and variables associated with the outcome were determined. Results: AKI was developed in 54.12% (n = 315) of patients, and these patients had blood pressure, SOFA score, APACHE II score, GCS, and various blood chemistry and hematology characteristics significantly different from the patients without AKI. Demographic characteristics (e.g. age and weight) were comparable between the two groups of patients. Among the 315 patients with AKI, 136 of them died during the study period. Multivariate logistic regression analysis revealed that the outcome of patients was associated with lung infection, coagulation system dysfunction, staphylococcus aureus infection, and use of various treatments (epinephrine, norepinephrine, and the use of mechanical ventilation) after AKI development. Conclusion: AKI occurred in approximately half of the critically ill patients admitted to ICU. The site and type of infections, as well as the use of vasopressor agents, were associated with the outcome. [ABSTRACT FROM AUTHOR]

[Display omitted] • Alternative strategy for the customized production of ω6-PUFAs was developed. • Optimal biosynthetic pathways for ω6-PUFAs in Y. lipolytica were investigated. • ω6-PUFAs production was improved by relieving the substrate competition bottleneck. • The highest proportion of DGLA at 46.65% of TFAs was achieved in Y. lipolytica. Omega-6 polyunsaturated fatty acids (ω6-PUFAs), such as γ-linolenic acid (GLA), dihomo-γ-linolenic acid (DGLA) and arachidonic acid (ARA), are indispensable nutrients for human health. Harnessing the lipogenesis pathway of Yarrowia lipolytica creates a potential platform for producing customized ω6-PUFAs. This study explored the optimal biosynthetic pathways for customized production of ω6-PUFAs in Y. lipolytica via either the Δ6 pathway from Mortierella alpina or the Δ8 pathway from Isochrysis galbana. Subsequently, the proportion of ω6-PUFAs in total fatty acids (TFAs) was effectively increased by bolstering the provision of precursors for fatty acid biosynthesis and carriers for fatty acid desaturation, as well as preventing fatty acid degradation. Finally, the proportions of GLA, DGLA and ARA synthesized by customized strains accounted for 22.58%, 46.65% and 11.30% of TFAs, and the corresponding titers reached 386.59, 832.00 and 191.76 mg/L in shake-flask fermentation, respectively. This work provides valuable insights into the production of functional ω6-PUFAs. [ABSTRACT FROM AUTHOR]

In recent years, eukaryotic microorganisms have been widely applied to offer many solutions for everyday life and have come to play important roles in agriculture, food, health care, and the fine-chemicals industry. However, the complex genetic background and low homologous recombination efficiency have hampered the implementation of large-scale and high-throughput gene editing in many eukaryotic microorganisms. The low efficiency of homologous recombination (HR) not only makes the modification process labor-intensive but also completely precludes the application of many otherwise very useful genome editing techniques. Thus, increasing the efficiency of HR is clearly an enabling technology for basic research and gene editing in eukaryotic microorganisms. In this review, we summarize the current strategies for enhancing the efficiency of HR in eukaryotic microorganisms (particularly yeasts and filamentous fungi), list some small molecules and candidate genes associated with homologous and non-homologous recombination, and briefly discuss the further development prospects of these strategies. [ABSTRACT FROM AUTHOR]

Highlights • Y. lipolytica is a promising chassis for the production of plant derived terpenoids. • Metabolic engineering strategies for terpenoids synthesis in Y. lipolytica are summarized. • Prospective for terpenoids synthesis in Y. lipolytica based on latest technology are discussed. Abstract Terpenoids are a large class of natural compounds based on the C5 isoprene unit, with many biological effects such activity against cancer and allergies, while some also have an agreeable aroma. Consequently, they have received extensive attention in the food, pharmaceutical and cosmetic fields. With the identification and analysis of the underlying natural product synthesis pathways, current microbial-based metabolic engineering approaches have yielded new strategies for the production of highly valuable terpenoids. Yarrowia lipolytica is a non-conventional oleaginous yeast that is rapidly emerging as a valuable host for the production of terpenoids due to its own endogenous mevalonate pathway and high oil production capacity. This review aims to summarize the status and strategies of metabolic engineering for the heterologous synthesis of terpenoids in Y. lipolytica in recent years and proposes new methods aiming towards further improvement of terpenoid production. [ABSTRACT FROM AUTHOR]

Abstract Microalgal lipids have drawn great attention as a promising sustainable resource for biodiesel or food supplement production. The development of high-performance strains of microalgae by metabolic engineering is invaluable for increasing the quantity or quality of desired lipids. The synthesis routes of lipids used as biodiesel in microalgae are based on fatty acid synthase (FAS) and triacylglycerols (TAG) biosynthesis pathway. Polyunsaturated fatty acids (PUFAs), including ω-6 and ω-3 fatty acids, are essential nutrients for humans. Notably, microalgae possess two distinct pathways for polyunsaturated fatty acids (PUFAs) biosynthesis, including the desaturase/elongase pathway and the polyketide synthase (PKS) pathway. Thus, it is necessary to identify which biosynthetic pathways are responsible for PUFA synthesis in particular microalgae species. In recent years, various key enzymes and functional domains involved in fatty acid and TAG biosynthesis pathway were identified and potentially regulated by genetic engineering approaches to elevate specific lipids content. In addition, other studies have reported the implementation of strategies to increase lipid accumulation based on increasing acetyl-CoA/NADPH supply, enhancing photosynthetic efficiency, or blocking competing pathways. Furthermore, other efforts have used transcription factor engineering to simultaneously regulate multiple genes related to lipid accumulation. This review summarizes recent research about a variety of microalgae lipid biosynthesis pathways, and discusses multiple gene manipulation strategies that have been employed for specific lipid overproduction in industrial microalgae. Highlights • Metabolic engineering of microalgae for producing biodiesel is reviewed. • Metabolic engineering of microalgae for producing food supplements is reviewed. • Two distinct polyunsaturated fatty acids biosynthesis pathway is discussed. • Regulation of lipid production by transcription factor engineering is summarized. • Further prospects on improvement microalgal lipids production is proposed. [ABSTRACT FROM AUTHOR]

Highlights • Algae-residue from Schizochytrium sp. was used as nitrogen source to replace yeast extract. • Cane molasses was used as carbon source to substitute glucose. • Two-stage culture with different carbon sources increased the efficiency of DHA production. • This work offers a strategy to reduce waste by-products discharge and cut production costs. Abstract The aim of this work was to reduce the algae-residue emission and make use of cane molasses as fermentation materials for docosahexaenoic acid (DHA) fermentaion by Schizochytrium sp., which further could cut the cost of DHA production. Algae-residue and cane molasses were respectively used as nitrogen and carbon sources to replace yeast extract and glucose. A significant DHA yield of 18.58 g/L was obtained using algae-residue, while cane molasses could not be used well as sole carbon source due to the presence of undesirable substance. A two-stage culture strategy with glucose followed by pretreated cane molasses as carbon source was developed, resulting in a final DHA yield of 15.22 g/L. This study therefore offers an economical and green strategy for DHA production by Schizochytrium sp. [ABSTRACT FROM AUTHOR]

The oleaginous yeast Yarrowia lipolytica is widely used for the production of both bulk and fine chemicals, including organic acids, fatty acid-derived biofuels and chemicals, polyunsaturated fatty acids, single-cell proteins, terpenoids, and other valuable products. Consequently, it is becoming increasingly popular for metabolic engineering applications. Multiple gene manipulation tools including URA blast, Cre/LoxP, and transcription activator-like effector nucleases (TALENs) have been developed for metabolic engineering in Y. lipolytica. However, the low efficiency and time-consuming procedures involved in these methods hamper further research. The emergence of the CRISPR/Cas system offers a potential solution for these problems due to its high efficiency, ease of operation, and time savings, which can significantly accelerate the genomic engineering of Y. lipolytica. In this review, we summarize the research progress on the development of CRISPR/Cas systems for Y. lipolytica, including Cas9 proteins and sgRNA expression strategies, as well as gene knock-out/knock-in and repression/activation applications. Finally, the most promising and tantalizing future prospects in this area are highlighted. [ABSTRACT FROM AUTHOR]

Lipid accumulation of Schizochytrium sp. can be induced by stress condition, but this stress-induction usually reduce cell growth and cause oxidative damage, which can eventually lower the lipid yield. Here, adaptive laboratory evolution (ALE) combined high salinity was performed to enhance the antioxidant system and lipid accumulation. The final strain ALE150, which was obtained after 150 days, showed a maximal cell dry weight (CDW) of 134.5 g/L and lipid yield of 80.14 g/L, representing a 32.7 and 53.31% increase over the starting strain, respectively. Moreover, ALE150 exhibited an overall higher total antioxidant capacity (T-AOC) and lower reactive oxygen species (ROS) levels than the starting strain. Furthermore, the regulatory mechanisms responsible for the improved performance of ALE150 were analyzed by transcriptomic analysis. Genes related to the antioxidant enzymes and central carbon metabolism were up-regulation. Moreover, the metabolic fluxes towards the fatty acid synthase (FAS) and polyketide synthase (PKS) pathways were also changed. [ABSTRACT FROM AUTHOR]

Fermentation wastewater (FW) and algal residue are major by-products of docosahexaenoic acid (DHA) fermentations utilizing Schizochytrium sp. In order to reduce production costs and environmental pollution, we explored the application of FW and algal-residue extract (AE) for DHA production. Components analysis showed that FW and AE contained some mineral elements and protein residues, respectively. When they were used for DHA fermentation, results showed that 20% replacement of fresh water by FW and 80% replacement of yeast extract nitrogen by AE reached DHA content of 22.23 g/L and 27.10 g/L, respectively. Furthermore, a novel medium that utilizes a mixture of FW and AE was applied for DHA fermentation, whereby the final DHA yield reached 28.45 g/L, 24.56% higher than conventional medium. The strategy of valorizing fermentation waste provides a new method for reducing the costs and reducing environmental pollution of microbial fermentations. [ABSTRACT FROM AUTHOR]

Metabolic profiling was used to discover mechanisms of increased pneumocandin B0 production in a high-yield strain by comparing it with its parent strain. Initially, 79 intracellular metabolites were identified, and the levels of 15 metabolites involved in six pathways were found to be directly correlated with pneumocandin B0 biosynthesis. Then by combining the analysis of key enzymes, acetyl-CoA and NADPH were identified as the main factors limiting pneumocandin B0 biosynthesis. Other metabolites, such as pyruvate, α-ketoglutaric acid, lactate, unsaturated fatty acids and previously unreported metabolite γ-aminobutyric acid were shown to play important roles in pneumocandin B0 biosynthesis and cell growth. Finally, the overall metabolic mechanism hypothesis was formulated and a rational feeding strategy was implemented that increased the pneumocandin B0 yield from 1821 to 2768 mg/L. These results provide practical and theoretical guidance for strain selection, medium optimization, and genetic engineering for pneumocandin B0 production. [ABSTRACT FROM AUTHOR]

The accumulation of reactive oxygen species (ROS) can induce oxidative damage which can be detrimental to microbial biomass and product yields. In this study, we used intermittent hypoxic stress (IHS) to eliminate the main source of ROS during the production of arachidonic acid (ARA)-rich lipids by Mortierella alpina . Biomass, lipid- and ARA concentration simultaneously reached their peak values, and the ARA yield increased by about 20% along with a lower level of ROS concentration. Under these conditions, more NADPH, substrate and energy may be available for lipid biosynthesis. The efficacy of this strategy was also confirmed in a bioreactor, with similar results. Interestingly, the total power required for stirring fell by about 4%, which may be of practical value for energy savings. This work offers new insights into the control of ROS generation in M. alpina and might be applicable to other microbial PUFAs producers. [ABSTRACT FROM AUTHOR]

[Display omitted] • Alternative strategy for nervonic acid-enriched plant oil production is developed. • Orthogonal plant and non-plant nervonic acid synthetic pathways are constructed. • MoLPAAT from Malania oleifera is identified to be specificity for nervonic acid. • The highest titer of nervonic acid at 13.56 g/L is achieved in Yarrowia lipolytica. Nervonic acid has proven efficacy in brain development and the prevention of neurodegenerative diseases. Here, an alternative and sustainable strategy for nervonic acid-enriched plant oil production was established. Different β-ketoacyl-CoA synthases and heterologous Δ15 desaturase were co-expressed, combined with the deletion of the β-oxidation pathway to construct orthogonal plant and non-plant nervonic acid biosynthesis pathways in Yarrowia lipolytica. A "block-pull-restrain" strategy was further applied to improve the supply of stearic acid as the precursor of the non-plant pathway. Then, lysophosphatidic acid acyltransferase from Malania oleifera (Mo Lpaat) was identified, which showed specificity for nervonic acid. Endogenous LPAAT was exchanged by MoLPAAT resulted in 17.10 % nervonic acid accumulation. Finally, lipid metabolism was engineered and cofactor supply was increased to boost the lipid accumulation in a stable null-hyphal strain. The final strain produced 57.84 g/L oils with 23.44 % nervonic acid in fed-batch fermentation, which has the potential to substitute nervonic acid-enriched plant oil. [ABSTRACT FROM AUTHOR]

[Display omitted] • Co-overexpression of GLK1 and PGM1 significantly promoted cordycepin accumulation. • Molasses, DHP, and waste yeast as alternate enhanced cordycepin production. • High C/N molar ratio and weak acidic were more conducive to producing cordycepin. • The cordycepin productivity in the optimized medium was increased by 28.81%. • Using agro-industrial residues can reduce the cost of cordycepin production. Cordycepin, a nucleoside compound with a variety of biological activities, has been extensively applied in the nutraceutical and pharmaceutical industries. The advancement of microbial cell factories using agro-industrial residues provides a sustainable pathway for cordycepin biosynthesis. Herein, the cordycepin production was enhanced by the modification of glycolysis and pentose phosphate pathway in engineered Yarrowia lipolytica. Then, cordycepin production based on economical and renewable substrates (sugarcane molasses, waste spent yeast, and diammonium hydrogen phosphate) was analyzed. Furthermore, the effects of C/N molar ratio and initial pH on cordycepin production were evaluated. Results indicated that the maximum cordycepin productivity of 656.27 mg/L/d (72 h) and cordycepin titer was 2286.04 mg/L (120 h) by engineered Y. lipolytica in the optimized medium, respectively. The cordycepin productivity in the optimized medium was increased by 28.81% compared with the original medium. This research establishes a promising way for efficient cordycepin production from agro-industrial residues. [ABSTRACT FROM AUTHOR]

Enhancing lipid productivity and reducing oxidative damage is essential for lipid overproduction in microalgae. In this study, addition of 20 mg/L fulvic acid (FA) resulted a 34.4% increase of lipid yield in Schizochytrium sp. Furthermore, the cooperative effect of FA and EDTA on cell growth and lipid production was investigated. The combined addition of 20 mg/L FA and 1.0 g/L EDTA yielded a maximal cell dry weight of 130.7 g/L and lipid productivity of 1.16 g/L/h, representing 36.4% and threefold increase over the non-supplemented group, respectively. Moreover, compared with the non-supplemented group, the combined addition strategy exhibited overall lower levels of reactive oxygen species and malondialdehyde, which accompanied with 66.7% and 81.9% higher superoxide dismutase and catalase activity, respectively. Furthermore, a 24.1-37.1% increase of malic enzyme and 19.4-25.2% decrease of phosphoenolpyruvate carboxylase activity was observed during the entire fermentation stage (0-108 h). Results suggested that the combined addition strategy not only enhanced lipid accumulation, but also prevented the lipid peroxidation. [ABSTRACT FROM AUTHOR]

The non‐conventional yeast Yarrowia lipolytica have been receiving growing attention due to its excellent lipid accumulation capacity. Microbial lipid have attracted widespread interest due to their broad applications as dietary supplements, cosmetic additives, oleochemicals, and renewable starting materials for the production of fossil fuel. With the development of whole‐genome sequencing, many effective genetic tools, including transformation systems, promoter systems, genomic integration and genome editing tools, have been applied in Y. lipolytica to enhance the overproduction of lipid. It can be genetically engineered for high lipid production via the upregulation of synthetic precursor and lipid synthesis pathways, the downregulation or disruption of competing pathways such as β‐oxidation, and elimination of inhibitory factors. In this review, the lipid metabolism, available genetic tools, and recent advances in metabolic engineering of Y. lipolytica for the overproduction of lipid and lipid‐derived chemicals is summarized. Future prospects of lipid biosynthesis in Y. lipolytica are discussed in light of the current progress, challenges, and trends in this field. Guidelines for future studies are also proposed. Practical Applications: General concerns about climate change, oil price crisis, and the increasing study for renewable energy are driving bio‐lipid as promising alternatives to fossil fuel. Over the past few decades, microbial lipid have been widely applied in dietary supplements, cosmetic additives, oleochemicals, and renewable starting materials for the production of fossil fuel. The non‐conventional yeast Yarrowia lipolytica have become an attractive metabolic engineering host for the production of microbial lipids due to its ability to synthesize them in large quantities. This review illuminates the lipid biosynthesis and degradation of Y. lipolytica, and summarizes the metabolic engineering efforts which have targeted a variety of biosynthetic biosynthetic pathway to efficiently convert carbon source to lipid in oleaginous Y. lipolytica. Schematic diagram of lipid and triacylglyceride biosynthesis, including the native and heterologous biosynthesis pathways of fatty acids in Y. lipolytica. [ABSTRACT FROM AUTHOR]

As one of the most important environmental factors, oxygen is particularly important for synthesis of n-3 polyunsaturated fatty acids (n-3 PUFA) in microalgae. In general, a higher oxygen supply is beneficial for cell growth but obstructs PUFA synthesis. The generation of reactive oxygen species (ROS) under aerobic conditions, which leads to the peroxidation of lipids and especially PUFA, is an inevitable aspect of life, but is often ignored in fermentation processes. Irritability, microalgal cells are able to activate a number of anti-oxidative defenses, and the lipid profile of many species is reported to be altered under oxidative stress. In this review, the effects of oxygen on the PUFA synthesis, sources of oxidative damage, and anti-oxidative defense systems of microalgae were summarized and discussed. Moreover, this review summarizes the published reports on microalgal biotechnology involving direct/indirect oxygen regulation and new bioreactor designs that enable the improved production of PUFA. [ABSTRACT FROM AUTHOR]

Synthetic multigene pathways enable microbes to synthesize novel products, but generally affect cell growth. In our previous study, an arachidonic acid biosynthesis pathway was constructed in Yarrowia lipolytica , and interestingly, the engineered strain achieved higher biomass production without metabolic burden. In this study, we provide an in-depth analysis of the effects of this multigene pathway on Y. lipolytica . Compared to the original strain, the engineered strain was able to produce arachidonic acid (0.42% of total fatty acids). Moreover, the engineered strain had robust growth and a low titer of extracellular organic acids. qRT-PCR analysis results indicated that the heterologous metabolic pathway influenced the expression of genes related to fatty acid metabolism and the secretion of organic acids. Taken together, it can thus be concluded that the synthetic metabolic pathway is able to redirect the carbon flux towards intracellular fatty acid accumulation at the expense of extracellular organic acid secretion in Y. lipolytica , which is highly desirable and can hopefully be copied in other strains. [ABSTRACT FROM AUTHOR]

In recent years, a variety of genetic tools have been developed and applied to various filamentous fungi, which are widely applied in agriculture and the food industry. However, the low efficiency of gene targeting has for many years hampered studies on functional genomics in this important group of microorganisms. The emergence of CRISPR/Cas9 genome-editing technology has sparked a revolution in genetic research due to its high efficiency, versatility, and easy operation and opened the door for the discovery and exploitation of many new natural products. Although the application of the CRISPR/Cas9 system in filamentous fungi is still in its infancy compared to its common use in E. coli, yeasts, and mammals, the deep development of this system will certainly drive the exploitation of fungal diversity. In this review, we summarize the research progress on CRISPR/Cas9 systems in filamentous fungi and finally highlight further prospects in this area. [ABSTRACT FROM AUTHOR]

To investigate the metabolic regulation against oxygen supply, comparative metabolomics was performed to explore the metabolic responses of Mortierella alpina in the process of arachidonic acid (ARA) production. More than 110 metabolites involved in Embden-Meyerhof-Parnas pathway, pentose phosphate pathway, tricarboxylic acid cycle, inositol phosphate metabolism, fatty acid biosynthesis, and amino acid metabolism were identified by gas chromatography-mass spectrometry. Samples at different aeration rates were clearly distinguished by principal components analysis and partial least squares analysis, indicating that oxygen supply had a profound effect on the metabolism of M. alpina. Eleven major metabolites were identified as potential biomarkers to be primarily responsible for the difference of metabolism. Further study of metabolic changes with the relevant pathways demonstrated that the levels of several intermediate metabolites in relation to central carbon metabolism changed remarkably via both processes and citrate and malate was supposed to play vital roles in polyunsaturated acid (PUFA) synthesis. Increase of myo-inositol and sorbitol were probably for osmo-regulation and redox balance, while enhanced phosphoric acid and pyroglutamic acid were supposed to have function in the activation of signal transduction pathway for stress resistance. The present study provides a novel insight into the metabolic responses of M. alpina to aeration rates and the metabolic characteristics during the ARA fermentation. [ABSTRACT FROM AUTHOR]

Yarrowia lipolytica , a non-conventional oleaginous yeast with special traits, has attracted increasing interest for producing value-added products. Generally, the DNA fragments of these heterologous metabolic pathways are constructed via the classic restriction digestion and ligation method. In contrast, the one-step in vivo pathway assembly method has been only rarely applied to Y. lipolytica . Here, with arachidonic acid biosynthesis as a case study, a one-step in vivo pathway assembly and integration method was used for engineering Y. lipolytica . Using rDNA as integrative locus, this study showed that there was a relation between the assembly efficiency and the length of overlapping region. Especially, with an overlap up to 1 kb, the method was able to rapidly assemble the arachidonic acid biosynthesis pathway (nearly 10 kb) into the chromosome with high efficiency (nearly 23%). Meanwhile, the pathway assembled in Y. lipolytica demonstrated long-term genetic stability and the engineered strain exhibited robust growth. Furthermore, this study demonstrated that the codon-optimized genes from Mortierella alpina can function efficiently in Y. lipolytica : a high level arachidonic acid production (0.4% of total fatty acids) was produced in the engineered strain. To our knowledge, this is the first time that this method is applied to Y. lipolytica for functional polyunsaturated fatty acids production. This method represents a powerful tool with potential for facilitating engineering applications in non-conventional yeasts. [ABSTRACT FROM AUTHOR]

γ-Linolenic acid (GLA) has various well-documented beneficial physiological effects and high biological significance. Because the natural supply of GLA is insufficient, microbial GLA production is a promising method for pharmaceutical and nutraceutical purposes. To establish and develop a biotechnological process for GLA production by Yarrowia lipolytica , the codon-optimized △6-desaturase from Mortierella alpina was introduced into this yeast under the control of the strong hp4d promoter. A recombinant Y. lipolytica strain was constructed, which produced 4.6% GLA in total fatty acids. By using a temperature-shift strategy of cultivation, consisting in preliminary growth at 28 °C followed by 6-day culture at 20 °C, optimal levels of dry cell weight (DCW), lipid content and GLA concentration were obtained from the recombinant strain: 18.55 g/L, 1.16 g/L and 71.6 mg/L, respectively. These DCW, lipid and GLA values were respectively 25.7%, 19.6% and 60.9% higher than those obtained in the control cultivation experiment at the standard constant temperature of 28 °C. This work demonstrates the excellent capacity of Y. lipolytica for GLA production, by combining metabolic engineering with a temperature-shift strategy. [ABSTRACT FROM AUTHOR]

Vitamin K (menaquinone or MK) plays an important role in blood clotting, cardiovascular disease, and anti-osteoporosis. A novel bacterial strain was isolated and identified as Bacillus natto based on 16SrDNA sequencing and LC-MS analysis. The objective of this study was to improve the extraction efficiency and productivity of MK-7 from B. natto. Acid-heating method efficiently disrupted B. natto cells for MK-7 extraction. Bacillus natto had a wide range of pH (5.0 ~ 9.0) for optimal growth. Its MK-7 yield was increased when rotation speed was increased to 200 rpm. The highest MK-7 yield was obtained when glycerol and soy peptone were used in the growth media. Batch fermentation was subsequently tested in 5 L bioreactor, which gave a high productivity of MK-7 (at 0.60 mg/L/h). A positive correlation between MK-7 yield and sporulation ratio was also found. This study provides valuable information on the extraction and production of menaquinone-7 from B. natto under submerged fermentation condition. [ABSTRACT FROM AUTHOR]

Unusual fatty acids with special carbon chain length or functional groups have been extensively used in the chemical, material, nutraceutical, and pharmaceutical industries. The traditional sources of these valuable fatty acids mainly include animal or plant extraction and chemical synthesis, which are unsustainable and may cause considerable environmental issues. The advancement of synthetic biology tools has facilitated the microbial production of lipids enriched in these fatty acids. The oleaginous yeast Yarrowia lipolytica is considered an attractive industrial host suitable for the production of advanced unusual fatty acids due to its high intrinsic lipogenesis ability. In this review, we introduce the most cutting-edge developments in synthetic biology tools for Y. lipolytica , as well as the recent progress in harnessing these tools to engineer the Y. lipolytica chassis to overaccumulate various unusual fatty acids, including odd-chain fatty acids, conjugated fatty acids, polyunsaturated fatty acids, cyclopropane fatty acids, methyl-branched fatty acids, hydroxylated fatty acids, and medium chain fatty acids. In addition, the future prospects of the unusual fatty acids using the Y. lipolytica platform are discussed in light of the current progress, challenges, and trends in this field. Finally, guidelines for future studies are also emphasized. [ABSTRACT FROM AUTHOR]