A Combined Manual Annotation and Deep-Learning Natural Language Processing Study on Accurate Entity Extraction in Hereditary Disease Related Biomedical Literature.

We report a combined manual annotation and deep-learning natural language processing study to make accurate entity extraction in hereditary disease related biomedical literature. A total of 400 full articles were manually annotated based on published guidelines by experienced genetic interpreters at Beijing Genomics Institute (BGI). The performance of our manual annotations was assessed by comparing our re-annotated results with those publicly available. The overall Jaccard index was calculated to be 0.866 for the four entity types-gene, variant, disease and species. Both a BERT-based large name entity recognition (NER) model and a DistilBERT-based simplified NER model were trained, validated and tested, respectively. Due to the limited manually annotated corpus, Such NER models were fine-tuned with two phases. The F1-scores of BERT-based NER for gene, variant, disease and species are 97.28%, 93.52%, 92.54% and 95.76%, respectively, while those of DistilBERT-based NER are 95.14%, 86.26%, 91.37% and 89.92%, respectively. Most importantly, the entity type of variant has been extracted by a large language model for the first time and a comparable F1-score with the state-of-the-art variant extraction model tmVar has been achieved.

Extraction methods and compositions of polyphenols in Shanxi aged vinegar.

The extraction, purification, qualification, and quantification of polyphenols (PPs) in vinegar are challenging owing to the complex matrix of vinegar and the specific physicochemical and structural properties of PPs. This study aimed to develop a simple, efficient, low-cost method for enriching and purifying vinegar PPs. The enrichment and purification effects of five solid phase extraction (SPE) columns and five macroporous adsorption resins (MARs) for PPs were compared. The results show that SPE columns were more effective in purifying vinegar PPs than MARs. Among them, the Strata-XA column showed a higher recovery (78.469 ± 0.949%), yield (80.808 ± 2.146%), and purity (86.629 ± 0.978%) than other columns. In total, 48 PPs were identified and quantified using SPE and gas chromatography-mass spectrometry from the SPE column extracts; phenolic acids, such as 4-hydroxyphenyllactic acid, vanillic acid, 4-hydroxycinnamic acid, 4-hydroxybenzoic acid, protocatechuic acid, and 3-(4-Hydroxy-3-methoxyphenyl) propionic acid, occupy a major position in SAV. Furthermore, considering the potential applications of PPs, the concentrates were characterized based on their bioactive properties. They exhibited high total PP, flavonoid, and melanoidin contents and excellent anti-glycosylation and antioxidant activities. These results indicate that the established methodology is a high-efficiency, rapid-extraction, and environment-friendly method for separating and purifying PPs, with broad application prospects in the food, chemical, and cosmetic industries.

Construction of automated high-throughput screening method for finding efficient 3-ketosteroid 1,2-dehydrogenating strains.

Dehydrogenation reaction at C1(2) positions is typical and representative of industrial production of steroid drugs. Anti-inflammatory activity can be doubled when the nucleus of the anti-inflammatory steroid hormone drug introduces double bonds at the C1(2) positions. Arthrobacter simplex is currently the most widely studied and used strain for C1(2) dehydrogenation. Therefore, breeding Arthrobacter simplex with high-efficiency dehydrogenation ability is of great significance. In order to obtain high-efficiency strains, the research proposed a new screening strategy based on image process technique: firstly, a color reaction between 2,4-dinitrophenylhydrazine (DNPH) and 9α-hydroxyandrost-4-ene-3,17-dione (9α-OH-AD) was established to characterize the dehydrogenation ability of the strain; secondly, the color data of strains mutated by atmospheric and room temperature plasma (ARTP) in the "color reaction" were automated and analyzed for dehydrogenation ability prediction using optimized support vector machine model. Result showed that the prediction accuracy reached as high as 96% in verification experiments. After a series of mutagenesis, including breaking the bottleneck of a single mutation in ARTP, the dominant strain ARLU-146 was finally obtained from 5168 strains. Its initial conversion rate was 0.8059 g/L/h, with a conversion of 94.41% at 24 h, compared to the original strain ASP which increased the transformation rate by more than 10%. By further process optimization, a high conversion (94.34% within 20 h) with high substrate (85 g/L cortisone acetate) was achieved. According to literature research, it is the highest conversion at this substrate concentration. KEY POINTS: • A high-throughput screening method was developed by using image processing and machine learning technique. • "Mutation bottleneck" of single ARTP mutagenesis was surpassed by complex mutagenesis. • A high substrate (85 g/L CA) and high transformation rate craft (94.34% within 20 h) were built.

Interaction of acetic acid bacteria and lactic acid bacteria in multispecies solid-state fermentation of traditional Chinese cereal vinegar.

The microbial community plays an important role on the solid-state fermentation (SSF) of Chinese cereal vinegar, where acetic acid bacteria (AAB) and lactic acid bacteria (LAB) are the dominant bacteria. In this study, the top-down (in situ) and bottom-up (in vitro) approaches were employed to reveal the interaction of AAB and LAB in SSF of Shanxi aged vinegar (SAV). The results of high-throughput sequencing indicates that Acetobacter pasteurianus and Lactobacillus helveticus are the predominant species of AAB and LAB, respectively, and they showed negative interrelationship during the fermentation. A. pasteurianus CGMCC 3089 and L. helveticus CGMCC 12062, both of which were isolated from fermentation of SAV, showed no nutritional competition when they were co-cultured in vitro. However, the growth and metabolism of L. helveticus CGMCC 12062 were inhibited during SSF due to the presence of A. pasteurianus CGMCC 3089, indicating an amensalism phenomenon between these two species. The transcriptomic results shows that there are 831 differentially expressed genes (|log2 (Fold Change)| > 1 and, p ≤ 0.05) in L. helveticus CGMCC 12062 under co-culture condition comparing to its mono-culture, which are mainly classified into Gene Ontology classification of molecular function, biological process, and cell composition. Of those 831 differentially expressed genes, 202 genes are up-regulated and 629 genes are down-regulated. The down-regulated genes were enriched in KEGG pathways of sugar, amino acid, purine, and pyrimidine metabolism. The transcriptomic results for A. pasteurianus CGMCC 3089 under co-culture condition reveals 529 differentially expressed genes with 393 up-regulated and 136 down-regulated, and the genes within KEGG pathways of sugar, amino acid, purine, and pyrimidine metabolism are up-regulated. Results indicate an amensalism relationship in co-culture of A. pasteurianus and L. helveticus. Therefore, this work gives a whole insight on the interaction between the predominant species in SSF of cereal vinegar from nutrient utilization, endogenous factors inhibition and the regulation of gene transcription.

Oxidoreduction potential controlling for increasing the fermentability of enzymatically hydrolyzed steam-exploded corn stover for butanol production.

BACKGROUND: Lignocellulosic biomass is recognized as an effective potential substrate for biobutanol production. Though many pretreatment and detoxification methods have been set up, the fermentability of detoxicated lignocellulosic substrate is still far lower than that of starchy feedstocks. On the other hand, the number of recent efforts on rational metabolic engineering approaches to increase butanol production in Clostridium strains is also quite limited, demonstrating the physiological complexity of solventogenic clostridia. In fact, the strain performance is greatly impacted by process control. developing efficient process control strategies could be a feasible solution to this problem. RESULTS: In this study, oxidoreduction potential (ORP) controlling was applied to increase the fermentability of enzymatically hydrolyzed steam-exploded corn stover (SECS) for butanol production. When ORP of detoxicated SECS was controlled at - 350 mV, the period of fermentation was shortened by 6 h with an increase of 27.5% in the total solvent (to 18.1 g/L) and 34.2% in butanol (to 10.2 g/L) respectively. Silico modeling revealed that the fluxes of NADPH, NADH and ATP strongly differed between the different scenarios. Quantitative analysis showed that intracellular concentrations of ATP, NADPH/NADP+, and NADH/NAD+ were increased by 25.1%, 81.8%, and 62.5%. ORP controlling also resulted in a 2.1-fold increase in butyraldehyde dehydrogenase, a 1.2-fold increase in butanol dehydrogenase and 29% increase in the cell integrity. CONCLUSION: ORP control strategy effectively changed the intracellular metabolic spectrum and significantly improved Clostridium cell growth and butanol production. The working mechanism can be summarized into three aspects: First, Glycolysis and TCA circulation pathways were strengthened through key nodes such as pyruvate carboxylase [EC: 6.4.1.1], which provided sufficient NADH and NADPH for the cell. Second, sufficient ATP was provided to avoid "acid crash". Third, the key enzymes activities regulating butanol biosynthesis and cell membrane integrity were improved.

Exploring polymerisation of methylglyoxal with NH3 or alanine to analyse the formation of typical polymers in melanoidins.

To investigate the formation of typical melanoidin polymers, methylglyoxal (MGO) with NH3 or alanine (Ala) was used to form coloured compounds, with glyoxal or acetone used as controls. The products were characterised using chromatography, mass spectrometry, and spectroscopy. Spectroscopic results showed that the coloured compounds formed were similar to melanoidins in food. GC-MS results showed that the MGO-based reaction generated similar volatile compounds using the Maillard reaction. Mass spectrometry showed that the molecular weights of structural units in the polymers were mainly 162, 169, and 176 Da, and these could be reassembled using the basic units derived from MGO alone or in combination with nitrogen. Hence, polymers recombined using basic structural units should be considered while determining melanoidin biomarkers. The preparation of coloured compounds using MGO with NH3 can be used as a novel method to produce the control compounds for melanoidin after process optimization.

Efficient One-Step Biocatalytic Multienzyme Cascade Strategy for Direct Conversion of Phytosterol to C-17-Hydroxylated Steroids.

Steroidal 17-carbonyl reduction is crucial to the production of natural bioactive steroid medicines, and boldenone (BD) is one of the important C-17-hydroxylated steroids. Although efforts have been made to produce BD through biotransformation, the challenges of the complex transformation process, high substrate costs, and low catalytic efficiencies have yet to be mastered. Phytosterol (PS) is the most widely accepted substrate for the production of steroid medicines due to its similar foundational structure and ubiquitous sources. 17ß-Hydroxysteroid dehydrogenase (17ßHSD) and its native electron donor play significant roles in the 17ß-carbonyl reduction reaction of steroids. In this study, we bridged 17ßHSD with a cofactor regeneration strategy in Mycobacterium neoaurum to establish a one-step biocatalytic carbonyl reduction strategy for the efficient biosynthesis of BD from PS for the first time. After investigating different intracellular electron transfer strategies, we rationally designed the engineered strain with the coexpression of 17ßhsd and the glucose-6-phosphate dehydrogenase (G6PDH) gene in M. neoaurum. With the establishment of an intracellular cofactor regeneration strategy, the ratio of [NADPH]/[NADP+] was maintained at a relatively high level, the yield of BD increased from 17% (in MNR M3M-ayr1S.c) to 78% (in MNR M3M-ayr1&g6p with glucose supplementation), and the productivity was increased by 6.5-fold. Furthermore, under optimal glucose supplementation conditions, the yield of BD reached 82%, which is the highest yield reported for transformation from PS in one step. This study demonstrated an excellent strategy for the production of many other valuable carbonyl reduction steroidal products from natural inexpensive raw materials. IMPORTANCE Steroid C-17-carbonyl reduction is one of the important transformations for the production of valuable steroidal medicines or intermediates for the further synthesis of steroidal medicines, but it remains a challenge through either chemical or biological synthesis. Phytosterol can be obtained from low-cost residues of waste natural materials, and it is preferred as the economical and applicable substrate for steroid medicine production by Mycobacterium. This study explored a green and efficient one-step biocatalytic carbonyl reduction strategy for the direct conversion of phytosterol to C-17-hydroxylated steroids by bridging 17ß-hydroxysteroid dehydrogenase with a cofactor regeneration strategy in Mycobacterium neoaurum. This work has practical value for the production of many valuable hydroxylated steroids from natural inexpensive raw materials.

Unraveling the metabolic network of organic acids in solid-state fermentation of Chinese cereal vinegar.

Shanxi aged vinegar (SAV) is fermented by multispecies microorganism with solid-state fermentation (SSF) technology, which contains a variety of organic acids. However, the metabolic network of them in SSF is still unclear. In this study, metagenomics technology was used to reveal the microbial community and functional genes in SAV fermentation. The metabolic network of key organic acids with taste active value higher than 1 was reconstructed for the first time, including acetate, lactate, malate, citrate, succinate, and tartrate. The results show pyruvate is the core compound in the metabolic network of organic acids. Metabolic pathway of acetate plays a pivotal role in this network, and acetate has regulatory function on metabolism of other organic acids. Acetobacter and Lactobacillus are the predominant genera for organic acid metabolism in SSF of SAV. This is also the first report on metabolic network of organic acids in cereal vinegar, adding new knowledge on the flavor substance metabolism during multispecies fermentation of traditional fermented food.

Metabolic profile of main organic acids and its regulatory mechanism in solid-state fermentation of Chinese cereal vinegar.

Shanxi aged vinegar (SAV), a traditional Chinese cereal vinegar, is produced using solid-state fermentation (SSF) technology. Organic acids are the key flavor compounds of vinegar. However, the metabolic mechanism of organic acids during SSF process is still unclear. In this study, metatranscriptomics was used to explore the metabolic profile of main organic acids in SSF. The results show that carbon metabolism is the dominant pathway during fermentation, among which pyruvate metabolism, glycolysis and starch and sucrose metabolism associated with organic acids were the most abundant. The metabolic pathways of acetic acid and lactic acid shift from acetyl-P and pyruvate pathways at early and middle-early stages of fermentation to acetaldehyde and L-lactaldehyde pathways at later stages, respectively, and Lactobacillus and Acetobacter are the predominant microorganisms contributed to them. Temperature and acetic acid are proven to be the environmental factors that regulate the metabolic activity during SSF. This study sheds new lights on metabolism of flavor substances in the spontaneous ecosystems of traditional fermented food.

Formation mechanisms and characterisation of the typical polymers in melanoidins from vinegar, coffee and model experiments.

Melanoidins, are of increasing interest for their potential biological activities. However, little knowledge is available on their structure. In the present study, vinegar, coffee and model melanoidins were degraded by NaBH4, and the resultant reaction products were characterised by chromatography, mass spectrometry and spectrometry methods to elucidate the mechanism of formation of melanoidin skeleton molecules. The study identified a typical polymer with a molecular weight (MW) interval of 74 Da, which was polymerised by aldol condensation and reduced by NaBH4, followed by intermolecular dehydration. MW of the theoretically derived typical polymers matched the detected polymers, validating the speculated pathway involved in the formation of melanoidins skeleton molecules. The study also revealed that melanoidins from different sources contain polymers with the same MW and different binding preferences, contributing to the heterogeneity of melanoidins. Overall, these findings indicated that the identified polymers could be used as potential candidate biomarkers for melanoidins.

Global regulator engineering enhances bioelectricity generation in Pseudomonas aeruginosa-inoculated MFCs.

The electricigens with high-electroactivity is essential for resolving the low electricity power output (EPT) of microbial fuel cells (MFCs). However, the manipulation by single functional genes shows limitation because electroactivity is a complex phenotype controlled by multiple genes. Herein, global regulator engineering (GRE) was developed to optimize the electroactivity of an isolated strain (Pseudomonas aeruginosa P3-A-11) using an exogenous global regulator IrrE (ionizing radiation resistance E linkage group) as an object. The GRE was implemented through in vitro random mutagenesis by error-prone PCR and in vivo high-through screening comprised of cultures color assay, PYO measurement and MFCs operation. Four mutants with higher electroactivity were obtained, among which, the mutant 11/M2-59 not only displayed the maximal power density, but also exhibited stronger salt tolerance, consequently showing good performance of MFCs in the presence of salt. Apart from the reduced internal resistance, the increase in phenazines amounts primarily contributed to EPT improvement, which was realized by enhancing the core biosynthesis pathway and affecting other pathways (such as central metabolism pathway, quorum sensing system, regulatory network). Notably, IrrE exerted its positive effect on electroactivity even without native regulators (such as PmpR and RpoS). In addition, the significant fluctuations in expression levels of stress-responsive genes mediated by GRE were closely associated with the enhanced salt tolerance. This work demonstrated that GRE was an effective approach for simultaneously optimizing multiple phenotypes (such as electroactivity and stress tolerance), and thus would provide more opportunities to create high-efficiency electricigens and further promoted the practical application of MFCs.

Development of optimal steam explosion pretreatment and highly effective cell factory for bioconversion of grain vinegar residue to butanol.

BACKGROUND: The industrial vinegar residue (VR) from solid-state fermentation, mainly cereals and their bran, will be a potential feedstock for future biofuels because of their low cost and easy availability. However, utilization of VR for butanol production has not been as much optimized as other sources of lignocellulose, which mainly stem from two key elements: (i) high biomass recalcitrance to enzymatic sugar release; (ii) lacking of suitable industrial biobutanol production strain. Though steam explosion has been proved effective for bio-refinery, few studies report SE for VR pretreatment. Much of the relevant knowledge remains unknown. Meanwhile, recent efforts on rational metabolic engineering approaches to increase butanol production in Clostridium strain are quite limited. In this study, we assessed the impact of SE pretreatment, enzymatic hydrolysis kinetics, overall sugar recovery and applied atmospheric and room temperature plasma (ARTP) mutant method for the Clostridium strain development to solve the long-standing problem. RESULTS: SE pretreatment was first performed. At the optimal condition, 29.47% of glucan, 71.62% of xylan and 22.21% of arabinan were depolymerized and obtained in the water extraction. In the sequential enzymatic hydrolysis process, enzymatic hydrolysis rate was increased by 13-fold compared to the VR without pretreatment and 19.60 g glucose, 15.21 g xylose and 5.63 g arabinose can be obtained after the two-step treatment from 100 g VR. Porous properties analysis indicated that steam explosion can effectively generate holes with diameter within 10-20 nm. Statistical analysis proved that enzymatic hydrolysis rate of VR followed the Pseudop-second-order kinetics equation and the relationship between SE severity and enzymatic hydrolysis rate can be well revealed by Boltzmann model. Finally, a superior inhibitor-tolerant strain, Clostridium acetobutylicum Tust-001, was generated with ARTP treatment. The water extraction and enzymolysis liquid gathered were successfully fermented, resulting in butanol titer of 7.98 g/L and 12.59 g/L of ABE. CONCLUSIONS: SE proved to be quite effective for VR due to high fermentable sugar recovery and enzymatic hydrolysate fermentability. Inverse strategy employing ARTP and repetitive domestication for strain breeding is quite feasible, providing us with a new tool for solving the problem in the biofuel fields.

The Sterol Carrier Hydroxypropyl-ß-Cyclodextrin Enhances the Metabolism of Phytosterols by Mycobacterium neoaurum.

Androst-4-ene-3,17-dione (AD) and androst-1,4-diene-3,17-dione (ADD) are valuable steroid pharmaceutical intermediates obtained by soybean phytosterol biotransformation by Mycobacterium Cyclodextrins (CDs) are generally believed to be carriers for phytosterol delivery and can improve the production of AD and ADD due to their effects on steroid solubilization and alteration in cell wall permeability for steroids. To better understand the mechanisms of CD promotion, we performed proteomic quantification of the effects of hydroxypropyl-ß-CD (HP-ß-CD) on phytosterol metabolism in Mycobacterium neoaurum TCCC 11978 C2. Perturbations are observed in steroid catabolism and glucose metabolism by adding HP-ß-CD in a phytosterol bioconversion system. AD and ADD, as metabolic products of phytosterol, are toxic to cells, with inhibited cell growth and biocatalytic activity. Treatment of mycobacteria with HP-ß-CD relieves the inhibitory effect of AD(D) on the electron transfer chain and cell growth. These results demonstrate the positive relationship between HP-ß-CD and phytosterol metabolism and give insight into the complex functions of CDs as mediators of the regulation of sterol metabolism.IMPORTANCE Phytosterols from soybean are low-cost by-products of soybean oil production and, owing to their good bioavailability in mycobacteria, are preferred as the substrates for steroid drug production via biotransformation by Mycobacterium However, the low level of production of steroid hormone drugs due to the low aqueous solubility (below 0.1 mmol/liter) of phytosterols limits the commercial use of sterol-transformed strains. To improve the bioconversion of steroids, cyclodextrins (CDs) are generally used as an effective carrier for the delivery of hydrophobic steroids to the bacterium. CDs improve the biotransformation of steroids due to their effects on steroid solubilization and alterations in cell wall permeability for steroids. However, studies have rarely reported the effects of CDs on cell metabolic pathways related to sterols. In this study, the effects of hydroxypropyl-ß-CD (HP-ß-CD) on the expression of enzymes related to steroid catabolic pathways in Mycobacterium neoaurum were systematically investigated. These findings will improve our understanding of the complex functions of CDs in the regulation of sterol metabolism and guide the application of CDs to sterol production.

Efficient repeated batch production of androstenedione using untreated cane molasses by Mycobacterium neoaurum driven by ATP futile cycle.

The biotransformation of phytosterol to androstenedione (AD) by mycobacteria is a unique process accompanied by energy-producing. However, high intracellular ATP content can severely inhibit the efficient production of AD. In this study, a novel citrate-based ATP futile cycle (AFC) and pyruvate-based AFC were constructed for the first time. Application of AFCs reduced intracellular ATP and propionyl-CoA levels and increased NAD+/NADH ratios and cell viability. The forced consumption of ATP promotes the transcription of critical genes in propionyl-CoA metabolism. The synergistic effect of enhanced propionyl-CoA metabolism and AFC increased AD conversion yield from 60.6% to 97.3%. The AD productivity was further improved by repeated batch fermentation using untreated cane molasses. The maximum productivity was 181% higher than that of the original strain. Therefore, the strategy of combining AFC and repeated batch fermentation is a valuable tool for the efficient and low-cost production of AD and other steroidal pharmaceutical precursors.

Isolation, characterisation, and genome sequencing of Rhodococcus equi: a novel strain producing chitin deacetylase.

Chitin deacetylase (CDA) can hydrolyse the acetamido group of chitin polymers to produce chitosans, which are used in various fields including the biomedical and pharmaceutical industries, food production, agriculture, and water treatment. CDA represents a more environmentally-friendly and easier to control alternative to the chemical methods currently utilised to produce chitosans from chitin; however, the majority of identified CDAs display activity toward low-molecular-weight oligomers and are essentially inactive toward polymeric chitin or chitosans. Therefore, it is important to identify novel CDAs with activity toward polymeric chitin and chitosans. In this study, we isolated the bacterium Rhodococcus equi F6 from a soil sample and showed that it expresses a novel CDA (ReCDA), whose activity toward 4-nitroacetanilide reached 19.20 U/mL/h during fermentation and was able to deacetylate polymeric chitin, colloidal chitin, glycol-chitin, and chitosan. Whole genome sequencing revealed that ReCDA is unique to the R. equi F6 genome, while phylogenetic analysis indicated that ReCDA is evolutionarily distant from other CDAs. In conclusion, ReCDA isolated from the R. equi F6 strain expands the known repertoire of CDAs and could be used to deacetylate polymeric chitosans and chitin in industrial applications.

Knowledge Domain and Emerging Trends in Vinegar Research: A Bibliometric Review of the Literature from WoSCC.

Vinegar is one of the most widely used acidic condiments. In recent decades, rapid advances have been made in the area of vinegar research, and the intellectual structure pertaining to this domain has significantly evolved. Thus, it is important that scientists keep abreast of associated developments to ensure an appropriate understanding of this field. To facilitate this current study, a bibliometric analysis method was adopted to visualize the knowledge map of vinegar research based on literature data retrieved from the Web of Science Core Collection (WoSCC) database. In total, 883 original research and review articles from between 1998 and 2019 with 19,663 references were analyzed by CiteSpace. Both a macroscopical sketch and microscopical characterization of the whole knowledge domain were realized. According to the research contents, the main themes that underlie vinegar research can be divided into six categories, that is, microorganisms, substances, health functions, production technologies, adjuvant medicines, and vinegar residues. In addition to the latter analysis, emerging trends and future research foci were predicted. Finally, the evolutionary stage of vinegar research was discerned according to Shneider's four-stage theory. This review will help scientists to discern the dynamic evolution of vinegar research, as well as highlight areas for future research.

Improving phytosterol biotransformation at low nitrogen levels by enhancing the methylcitrate cycle with transcriptional regulators PrpR and GlnR of Mycobacterium neoaurum.

BACKGROUND: Androstenedione (AD) is an important steroid medicine intermediate that is obtained via the degradation of phytosterols by mycobacteria. The production process of AD is mainly the degradation of the phytosterol aliphatic side chain, which is accompanied by the production of propionyl CoA. Excessive accumulation of intracellular propionyl-CoA produces a toxic effect in mycobacteria, which restricts the improvement of production efficiency. The 2-methylcitrate cycle pathway (MCC) plays a significant role in the detoxification of propionyl-CoA in bacterial. The effect of the MCC on phytosterol biotransformation in mycobacteria has not been elucidated in detail. Meanwhile, reducing fermentation cost has always been an important issue to be solved in the optimizing of the bioprocess. RESULTS: There is a complete MCC in Mycobacterium neoaurum (MNR), prpC, prpD and prpB in the prp operon encode methylcitrate synthase, methylcitrate dehydratase and methylisocitrate lyase involved in MCC, and PrpR is a specific transcriptional activator of prp operon. After the overexpression of prpDCB and prpR in MNR, the significantly improved transcription levels of prpC, prpD and prpB were observed. The highest conversion ratios of AD obtained by MNR-prpDBC and MNR-prpR increased from 72.3 ± 2.5% to 82.2 ± 2.2% and 90.6 ± 2.6%, respectively. Through enhanced the PrpR of MNR, the in intracellular propionyl-CoA levels decreased by 43 ± 3%, and the cell viability improved by 22 ± 1% compared to MNR at 96 h. The nitrogen transcription regulator GlnR repressed prp operon transcription in a nitrogen-limited medium. The glnR deletion enhanced the transcription level of prpDBC and the biotransformation ability of MNR. MNR-prpR/ΔglnR was constructed by the overexpression of prpR in the glnR-deleted strain showed adaptability to low nitrogen. The highest AD conversion ratio by MNR-prpR/ΔglnR was 92.8 ± 2.7% at low nitrogen level, which was 1.4 times higher than that of MNR. CONCLUSION: Improvement in phytosterol biotransformation after the enhancement of propionyl-CoA metabolism through the combined modifications of the prp operon and glnR of mycobacteria was investigated for the first time. The overexpress of prpR in MNR can increase the transcription of essential genes (prpC, prpD and prpB) of MCC, reduce the intracellular propionyl-CoA level and improve bacterial viability. The knockout of glnR can enhance the adaptability of MNR to the nitrogen source. In the MNRΔglnR strain, overexpress of prpR can achieve efficient production of AD at low nitrogen levels, thus reducing the production cost. This strategy provides a reference for the economic and effective production of other valuable steroid metabolites from phytosterol in the pharmaceutical industry.

Bioaugmentation by Pediococcus acidilactici AAF1-5 Improves the Bacterial Activity and Diversity of Cereal Vinegar Under Solid-State Fermentation.

Bioaugmentation technology may be an effective strategy to improve the solid-state fermentation rate and utilization of raw materials for traditional vinegar production. The relationship between bacteria and fermentation process was analyzed to rationally design and perform bioaugmented solid-state fermentation of the Tianjin Duliu mature vinegar (TDMV). Fermentation process was highly correlated with Acetobacter, Lactobacillus, and Pediococcus contents, which were the core functional microorganisms in TDMV fermentation. Pediococcus acidilactici AAF1-5 was selected from 20 strains to fortify the fermentation due to its acidity and thermal tolerance. Bioaugmentation was performed in the upper layer of TDMV fermentation. P. acidilactici AAF1-5 colonized and then spread into the lower layer to improve the fermentation. Result showed that the fermentation period was 5 days less than that of the control. Meanwhile, the non-volatile acid, lactic acid, amino nitrogen, and reducing sugar contents in the bioaugmented TDMV increased by 53%, 14%, 32%, and 36%, respectively, compared with those in the control. Bioaugmentation with P. acidilactici AAF1-5 not only improved the utilization of starch from 79% to 83% but also increased the bacterial community diversity.

Effects of Organic Acids, Amino Acids and Phenolic Compounds on Antioxidant Characteristic of Zhenjiang Aromatic Vinegar.

Zhenjiang aromatic vinegar (ZAV) is one of the famous Chinese vinegars, which contains various physicochemical and bioactive compositions. In the present study, physicochemical properties and total antioxidant activity were detected in ZAV samples. The correlation between of organic acids, amino acids, phenolic compounds, and the antioxidant activity of ZAV were explored. The results showed that contents of total acids, soluble solids, reducing sugar and total antioxidant activity in ZAV were increased with aging time, and those in ZAV-5 were the highest. Organic acids and amino acids exhibited weak antioxidant activity, while phenolic compounds had higher antioxidant ability. In addition, amino acids had synergistic effect on the antioxidant activity of phenolic compounds, whereas organic acids inhibited the antioxidant activity of phenolic compounds. Moreover, it was found that phenolic compounds including catechin, vanillic acid and syringic acid showed higher contribution rates to antioxidant activities of mixed phenolic compounds. In conclusion, these findings would provide references to control the antioxidant characteristic of vinegar through regulating the main compositions, and further improve the quality of vinegar production.

Economical production of androstenedione and 9α-hydroxyandrostenedione using untreated cane molasses by recombinant mycobacteria.

Production of androstenedione (AD) and 9α-hydroxyandrostenedione (9α-OH-AD) by recombinant mycobacteria using untreated cane molasses and hydrolysate of mycobacterial cells (HMC) was investigated for the first time. B-vitamins feeding experiment and reverse transcription-PCR analysis showed that propionyl-CoA carboxylase (PCC) plays an important role in the phytosterol biotransformation of mycobacteria. The respective AD and 9α-OH-AD conversion ratios were increased by 2.91 and 1.48 times through coexpression of PCC and NADH dehydrogenase. The highest conversion ratios of AD and 9α-OH-AD obtained by using a co-feeding strategy of cane molasses and HMC reached 96.38% and 95.04%, respectively, and the total costs of carbon and nitrogen sources for the culture medium were reduced by 29.89% and 49.49%, respectively. Taking the results together, untreated cane molasses and HMC can be used for the economical production of steroidal pharmaceutical precursors by mycobacteria. This study offers an economical and green strategy for steroidal pharmaceutical precursor production.