Whole genome sequencing and analysis of Bacillus sp. TTMP2, a tetramethylpyrazine-producing bacterium.

BACKGROUND: Tetramethylpyrazine has been extensively studied as an anticancer substance and a flavor substance in the fields of medicine and food industry. A strain with high tetramethylpyrazine production was screened from the fermented grains of Danquan winery. Genome sequencing can reveal the potential roles of bacteria by thoroughly examining the connection between genes and phenotypes from a genomic perspective. METHODS AND RESULTS: In this study, whole genome of this strain was sequenced and analyzed. This paper summarized the genomic characteristics of strain TTMP2 and analyzed genes related to the synthesis of tetramethylpyrazine. Bacillus sp. TTMP2 has a complete metabolic pathway for acetoin and tetramethylpyrazine metabolism. Gene function was analyzed by COG annotation, GO annotation, KEGG annotation and functional annotations for lipoproteins, carbohydrate-active enzymes, and pathogen-host interactions. Phylogenetic analysis indicated that Bacillus velezensis had the high homology with Bacillus sp. TTMP2. Genomes of 16 Bacillus species cover all genes of Bacillus, suggesting that genus Bacillus has an open pan-genome and can survive in diverse environments. CONCLUSION: The analysis of genome sequencing data from Bacillus sp. TTMP2 showed that its metabolic characteristics could be deeply understood, indicating that this bacterium had a particular role in tetramethylpyrazine synthesis.

Production of Tetramethylpyrazine from Cane Molasses by Bacillus sp. TTMP20.

2,3,5,6-Tetramethylpyrazine (TTMP) is an active ingredient of Ligusticum wallichii Franch. It can be used in medicine and food fields. In this study, Bacillus sp. TTMP20 was applied to produce TTMP using cane molasses as a carbon source. After pretreatment with phosphoric acid, 170 mL/L treated molasses, combined with 10 g/L yeast powder, 30 g/L tryptone and 30 g/L (NH4)2HPO4 were used for fermentation. After 36 h, TTMP output reached the highest value of 208.8 mg/L. The yield of TTMP using phosphoric acid-treated molasses as carbon source was 145.59% higher than control. Under the sulfuric acid treatment process of molasses (150 g), the maximum yield of TTMP was 895.13 mg/L, which was 183.18% higher than that of untreated molasses (316.1 mg/L). This study demonstrated that molasses is a high-quality and inexpensive carbon source for the manufacture of TTMP, laying the groundwork for the future industrial production of TTMP.

Optimization of Molasses and Soybean Meal Content to Enhance Tetramethylpyrazine Yield by Bacillus sp. TTMP20.

Microbial fermentation for the production of tetramethylpyrazine (TTMP) is considered to be the most promising method, and the development of a cheap fermentation substrate is of great importance for large-scale TTMP production. In this study, inexpensive by-products from the food industry, i.e., molasses and soybean meal (instead of glucose and tryptone), were used as substrates for TTMP fermentation. The pretreatment of soybean meal was explored in order to achieve a better fermentation effect. The contents of each component in the fermentation medium were optimized by central composite design (CCD). The optimum contents were as follows: 72.5 g/L of molasses, 37.4 g/L of diammonium hydrogen phosphate (DAP), 53.4 g/L of soybean meal, and 5 g/L of yeast powder. The software predicted a maximum TTMP yield of 1469.03 mg/L, and the actual TTMP yield was 1328.95 mg/L for the validation experiment in the optimum medium. Under the optimum conditions (72.5 g/L of molasses, 37.4 g/L of DAP, 53.4 g/L of soybean meal, and 5 g/L of yeast powder), the actual maximum TTMP yield (1328.95 mg/L) in this study was much higher than the TTMP yield (895.13 mg/L) under the conditions (150 g/L of molasses, 30 g/L of DAP, 30 g/L of tryptone, and 10 g/L of yeast powder) of our previous study published in Molecules. In this study, the TTMP yield improved by 48.46%, with decreased molasses (more than half), decreased yeast powder (half) and by-product soybean meal instead of tryptone compared to our previous study. In summary, the cheaper fermentation medium had a higher TTMP yield in this study, which improves the application potential of Bacillus sp. TTMP20.

Optimization of carotenoid extraction of a halophilic microalgae.

Dunaliella parva can produce abundant carotenoids under certain conditions. This paper optimized the extraction efficiency of carotenoids from D. parva. Different organic solvents were examined to determine the most suitable solvent for the extraction. After the determination of the solvent (dimethylsulfoxide, DMSO), the extraction conditions including time, temperature, and volume were then optimized to maximize the extraction efficiency of carotenoids from D. parva using response surface methodology. DMSO was identified as the most suitable solvent. The optimal extraction conditions were as follows: temperature of 57.2°C, time of 11.35 min, the volume of 410 µl, and the optimal extraction efficiency reached 0.517‰. The results showed that the optimal extraction efficiency (0.517‰) improved 31.69% in comparison to the initial extraction efficiency (0.3926‰). In addition, The optimal levels of three influence factors (temperature of 57.2°C, time of 11.35 min, volume of 410 µl) decreased compared with the initial levels (temperature of 60°C, time of 20 min, volume of 1000 µl). In this paper, Central Composite Design (CCD) was used to optimize the extraction efficiency of carotenoids from D. parva, which would lay the groundwork for the extraction and utilization of carotenoids from D. parva in the future.

Analysis of Bacterial Diversity in Different Types of Daqu and Fermented Grains From Danquan Distillery.

Bacterial communities in high-temperature Daqu and fermented grains are important for brewing Jiang-flavor Baijiu such as Danquan Baijiu. Daqu is a saccharifying and fermenting agent, which has a significant impact on the flavor of Baijiu. However, bacterial communities in three different types of samples from the Danquan distillery (dqjq_ck, dqjqcp, and dqjp3) were still unclear, which limited further development of Danquan Baijiu. "dqjq_ck" and "dqjqcp" indicate high-temperature Daqu at days 45 and 135, respectively. "dqjp3" indicates fermented grains. In this study, the bacterial communities of three samples were analyzed by Illumina Miseq high-throughput sequencing. The bacterial communities of three samples primarily composed of thermophilic bacteria and bacteria with stress resistance. The most abundant species in dqjq_ck, dqjqcp, and dqjp3 were Comamonas, Bacillus, and unclassified Lactobacillales, respectively. The main bacteria included Bacillus, Comamonas, Myroides, Paenibacillus, Acetobacter, Kroppenstedtia, Staphylococcus, Saccharopolyspora, Planifilum, Lactobacillus, Acinetobacter, Oceanobacillus, Enterococcus, Thermoactinomyces, Lactococcus, Streptomyces, Saccharomonospora, Tepidimicrobium, Anaerosalibacter, unclassified_Lactobacillales, unclassified_Thermoactinomycetaceae_1, unclassified_Bacillaceae_2, unclassified_Bacillales, unclassified_Microbacteriaceae, unclassified_Rhodobacteraceae, unclassified_Actinopolysporineae, and unclassified_Flavobacteriaceae in three samples (percentage was more than 1% in one of three samples). In our study, the succession of microbiota in three samples representing three important stages of Danquan Baijiu brewing was revealed. This article lays a good foundation for understanding the fermentation mechanism and screening some excellent indigenous bacteria to improve the quality of Danquan Baijiu in future.

Advanced glycosylated end products restrain the osteogenic differentiation of the periodontal ligament stem cell.

BACKGROUND/PURPOSE: Many studies have confirmed that periodontal disease interacts with diabetes. The aim of this study was to examine whether the advanced glycosylated end products (AGEs), which are generated by diabetics, have important effects on the osteogenic differentiation of periodontal ligament stem cells (PDLSCs). MATERIALS AND METHODS: In this study PDLSCs were isolated from the periodontal ligaments of extracted third molar teeth. The subjects were divided into two groups, which included the normal control group (N-PDLSCs) and the AGEs-stimulating group (A-PDLSCs). Changes of receptor of AGEs (RAGE) and cumulative ROS in PDLSCs were monitored by western blot and flow cytometry, respectively. RESULTS: In the study AGEs noticeably inhibited the osteogenic differentiation of PDLSCs, with significant lower calcification nodules detected in A-PDLSCs (P < 0.01). RAGE expression level and ROS accumulation in A-PDLSCs were clearly higher than those in N-PDLSCs (P < 0.01). CONCLUSION: Our conclusions were that AGEs may cause the apoptosis of stem cells, which could lead to the disorder of bone differentiation function of PDLSCs.