Optimization of Pre-treatment Using RSM on Wheat Straw and Production of Lactic Acid Using Thermotolerant, Inhibitor Tolerant and Xylose Utilizing Bacillus Sonorenesis Strain DGS15

Thermotolerant lactic acid producing bacteria, isolated from red soil of brick kiln was identified by 16S rRNA sequencing as Bacillus sonorenesis , which showed remarkable capability to ferment sugars of lignocellulosic biomass after pre-treatment, yielding 0.97 g/g lactic acid with overall productivity of 0.38 g L -1/ h. RSM was employed to optimize the sulphuric acid pre-treatment combined with dilute NaOH and hot water pre-treatment. Pretreated wheat straw biomass had 40.4% cellulose, 18.4% hemicellulose, 12.4% lignin and 28.2 g L -1 reducing sugar, while native wheat straw biomass had 36% cellulose, 25% hemicellulose, 20% total lignin, and 0.94 g L -1 reducing sugar. Scanning electron microscopy (SEM) revealed that the ordered and compact structure of wheat straw was destroyed upon pre-treatment. X-ray diffractogram (XRD) revealed 9.71% increase in crystallinity index ( CrI ) in pretreated biomass. FTIR spectrogram showed removal of lignin due to reduction of peak at 1640 cm -1 in pretreated biomass. Bacillus sonorenesis DGS15 is inhibitor tolerant (furfural (1.2 g L -1 ) and HMF (2.4 g L -1 )). Furfural was consumed after 72 h of fermentation and HMF got accumulated with 3.75-fold increase in concentration in the fermentation broth. In terms of final concentration, yield, and fermentation duration, this is the best performance of DGS15 for lactic acid production utilizing xylose, glucose as the carbon source. All of these findings showed that the thermotolerant Bacillus sonorenesis strain DGS15 is a novel, attractive candidate for producing lactic acid from lignocellulosic biomass.

Whole-Genome Sequencing and Potassium-Solubilizing Mechanism of Bacillus aryabhattai SK1-7

To analyze the whole genome of Bacillus aryabhattai strain SK1-7 and explore its potassium solubilization characteristics and mechanism, thus providing a theoretical basis for analyzing the utilization and improvement of insoluble potassium resources in soil. Genome information for Bacillus aryabhattai SK1-7 was obtained by using Illumina NovaSeq second-generation sequencing and GridION Nanopore ONT third-generation sequencing technology. The contents of organic acids and polysaccharides in fermentation broth of Bacillus aryabhattai SK1-7 were determined by high-performance liquid chromatography and the anthrone sulfuric acid method, and the expression levels of the potassium solubilization-related genes ackA, epsB, gltA, mdh and ppc were compared by real-time fluorescence quantitative PCR under different potassium source culture conditions. The whole genome of the strain consisted of a complete chromosome sequence and four plasmid sequences. The sequence sizes of the chromosomes and plasmids P1, P2, P3 and P4 were 5,188,391 bp, 136,204 bp, 124,862 bp, 67,200 bp and 12,374 bp, respectively. The GC contents were 38.2, 34.4, 33.6, 32.8, and 33.7%. Strain SK1-7 mainly secreted malic, formic, acetic and citric acids under culture with an insoluble potassium source. The polysaccharide content produced with an insoluble potassium source was higher than that with a soluble potassium source. The expression levels of five potassium solubilization-related genes with the insoluble potassium source were higher than those with the soluble potassium source.

Novel Millet-Based Flavored Yogurt Enriched With Superoxide Dismutase

Superoxide dismutase (SOD) is an important antioxidant enzyme with different physiological functions, which can be used as a nutritional fortifier in food. Cereal-based fermented products are becoming popular worldwide. In this study, novel millet-based flavored yogurt enriched with SOD was developed. Lactiplantibacillus plantarum subsp. plantarum was screened, which manufactured SOD activity of 2476.21 ± 1.52 U g−1. The SOD content of millet yogurt was 19.827 ± 0.323 U mL−1, which was 63.01, 50.11, and 146.79% higher than that of Bright Dairy Yogurt 1911, Junlebao and Nanjing Weigang, respectively. Fifty-four volatile flavor substances and 22,571 non-volatile flavor substances were found in yogurt. Compared to traditional fermented yogurt, 37 non-volatile metabolites in yogurt with millet enzymatic fermentation broth were significantly upregulated, including 2-phenyl ethanol, hesperidin, N-acetylornithine and L-methionine, which were upregulated by 3169.6, 228.36, 271.22, and 55.67 times, respectively, thereby enriching the sensory and nutritional value of yogurt. Moreover, the manufacture of unpleasant volatile flavor substances was masked, making the product more compatible with consumers' tastes.

Endophytic fungus Pseudodidymocyrtis lobariellae KL27 promotes taxol biosynthesis and accumulation in Taxus chinensis

Background Taxol from Taxus species is a precious drug used for the treatment of cancer and can effectively inhibit the proliferation of cancer cells. However, the growth of Taxus plants is very slow and the content of taxol is quite low. Therefore, it is of great significance to improve the yield of taxol by modern biotechnology without destroying the wild forest resources. Endophytic fungus which symbiosis with their host plants can promote the growth and secondary metabolism of medicinal plants. Results Here, an endophytic fungus KL27 was isolated from T. chinensis, and identified as Pseudodidymocyrtis lobariellae. The fermentation broth of KL27 (KL27-FB) could significantly promote the accumulation of taxol in needles of T. chinensis, reaching 0.361 ± 0.082 mg/g·DW (dry weight) at 7 days after KL27-FB treatment, which is 3.26-fold increase as compared to the control. The RNA-seq and qRT-PCR showed that KL27-FB could significantly increase the expression of key genes involved in the upstream pathway of terpene synthesis (such as DXS and DXR) and those in the taxol biosynthesis pathway (such as GGPPS, TS, T5OH, TAT, T10OH, T14OH, T2OH, TBT, DBAT and PAM), especially at the early stage of the stimulation. Moreover, the activation of jasmonic acid (JA) biosynthesis and JA signal transduction, and its crosstalk with other hormones, such as gibberellin acid (GA), ethylene (ET) and salicylic acid (SA), explained the elevation of most of the differential expressed genes related to taxol biosynthesis pathway. Moreover, TF (transcriptional factor)-encoding genes, including MYBs, ethylene-responsive transcription factors (ERFs) and basic/helix-loop-helix (bHLH), were detected as differential expressed genes after KL27-FB treatment, further suggested that the regulation of hormone signaling on genes of taxol biosynthesis was mediated by TFs. Conclusions Our results indicated that fermentation broth of endophytic fungus KL27-FB could effectively enhance the accumulation of taxol in T. chinensis needles by regulating the phytohormone metabolism and signal transduction and further up-regulating the expression of multiple key genes involved in taxol biosynthesis. This study provides new insight into the regulatory mechanism of how endophytic fungus promotes the production and accumulation of taxol in Taxus sp.

Supplementing Glycerol to Inoculum Induces Changes in pH, SCFA Profiles and Microbiota Composition in In-Vitro Batch Fermentation

Glycerol was generally added to the inoculum as a cryoprotectant. However, it was also a suitable substrate for microbial fermentation, which may produce more SCFAs, thereby decreased pH of the fermentation broth. This study investigated the effect of supplementing glycerol to inoculum on in vitro fermentation and whether an enhanced buffer capacity of medium could maintain the pH stability during in vitro batch fermentation, subsequently improving the accuracy of short chain fatty acids (SCFAs) determination, especially propionate. Two ileal digesta were fermented by pig fecal inoculum with or without glycerol (served as anti-frozen inoculum or frozen inoculum) in standard buffer or enhanced buffer solution (served as normal or modified medium). Along with the fermentation, adding glycerol decreased the pH of fermentation broth (p < 0.05). However, modified medium could alleviate the pH decrement compared with normal medium (p < 0.05). The concentration of total propionic acid production was much higher than that of other SCFAs in anti-frozen inoculum fermentation at 24 and 36 h, thereby increasing the variation (SD) of net production of propionate. The α-diversity analysis showed that adding glycerol decreased Chao1 and Shannon index under normal medium fermentation (p < 0.05) compared to modified medium (p < 0.05) along with fermentation. PCoA showed that all groups were clustered differently (p < 0.01). Adding glycerol improved the relative abundances of Firmicutes, Anaerovibrio, unclassified_f_Selenomonadaceae, and decreased the relative abundance of Proteobacteria (p < 0.05). The relative abundances of Firmicutes, such as Lactobacillus, Blautia and Eubacterium_Ruminantium_group in modified medium with frozen inoculum fermentation were higher than (p < 0.05) those in normal medium at 36 h of incubation. These results showed that adding glycerol in inoculum changed the fermentation patterns, regardless of substrate and medium, and suggested fermentation using frozen inoculum with modified medium could maintain stability of pH, improve the accuracy of SCFA determination, as well as maintain a balanced microbial community.

Sweet sorghum: broth clarification with enzymatic treatment increases the quality of the fermentation wort for ethanol production

Sweet sorghum is currently being evaluated throughout the world as a raw material for biofuel production because its stem juices are rich in sugars that can be directly fermented to ethanol. In this work, the fermentative efficiency of three sweet sorghum genotypes was evaluated, aiming at ethanol production, harvested in two seasons, clean and whole stems, and the treatment of the juice and broth with amylolytic enzymes in order to use the present starch to increase the production of ethanol. The experiment was carried out in the 2013/2014 harvest, in the municipality of Jaboticabal, São Paulo, Brasil, located at 21°14’05’’S and 48°17’09’’W. The experimental design was completely randomized, with sub-subdivided plots and four replications. The primary treatments were the sweet sorghum genotypes (CV147, CV198, and BRS508), the secondary treatments, the type of harvest (whole stems and clean stems); the tertiary the two sampling times (102 and 116 days after sowing - d.a.s) and the quaternary the application of enzymes. In the fermentation process, the yeast PE-2 was used, at the end, the wine was recovered and characterized. Fermentation efficiency and liters of ethanol per ton of sorghum were calculated. The clarification of the juice with enzymatic treatment increases the quality of the fermentation broth and makes it possible to obtain wines with lower levels of RRTs and Brix. Fermentation efficiency is not affected by the genotype; however, it is influenced by the time of harvest and the technological quality of the juice. The use of amylolytic enzymes makes it possible to obtain wines with lower levels of RRTS and Brix. The best period of industrialization was at 102 d.a.s., and the processing of whole stalks resulted in less ethanol production.

Efficient Extraction of Fermentation Inhibitors by Means of Green Hydrophobic Deep Eutectic Solvents

The methods for hydrogen yield efficiency improvements, the gaseous stream purification in gaseous biofuels generation, and the biomass pretreatment are considered as the main trends in research devoted to gaseous biofuel production. The environmental aspect related to the liquid stream purification arises. Moreover, the management of post-fermentation broth with the application of various biorefining techniques gains importance. Chemical compounds occurring in the exhausted liquid phase after biomass pretreatment and subsequent dark and photo fermentation processes are considered as value-added by products. The most valuable are furfural (FF), 5-hydroxymethylfurfural (HMF), and levulinic acid (LA). Enriching their solutions can be carried with the application of liquid–liquid extraction with the use of a suitable solvent. In these studies, hydrophobic deep eutectic solvents (DESs) were tested as extractants. The screening of 56 DESs was carried out using the Conductor-like Screening Model for Real Solvents (COSMO-RS). DESs which exposed the highest inhibitory effect on fermentation and negligible water solubility were prepared. The LA, FF, and HMF were analyzed using FT-IR and NMR spectroscopy. In addition, the basic physicochemical properties of DES were carefully studied. In the second part of the paper, deep eutectic solvents were used for the extraction of FF, LA, and HMF from post-fermentation broth (PFB). The main extraction parameters, i.e., temperature, pH, and DES: PFB volume ratio (VDES:VPFB), were optimized by means of a Box–Behnken design model. Two approaches have been proposed for extraction process. In the first approach, DES was used as a solvent. In the second, one of the DES components was added to the sample, and DES was generated in situ. To enhance the post-fermentation broth management, optimization of the parameters promoting HMF, FF, and LA extraction was carried under real conditions. Moreover, the antimicrobial effect of the extraction of FF, HMF, and LA was investigated to define the possibility of simultaneous separation of microbial parts and denatured peptides via precipitation.

Volatile Organic Compounds from Bacillus aryabhattai MCCC 1K02966 with Multiple Modes against Meloidogyne incognita

Plant-parasitic nematodes cause severe losses to crop production and economies all over the world. Bacillus aryabhattai MCCC 1K02966, a deep-sea bacterium, was obtained from the Southwest Indian Ocean and showed nematicidal and fumigant activities against Meloidogyne incognita in vitro. The nematicidal volatile organic compounds (VOCs) from the fermentation broth of B. aryabhattai MCCC 1K02966 were investigated further using solid-phase microextraction gas chromatography-mass spectrometry. Four VOCs, namely, pentane, 1-butanol, methyl thioacetate, and dimethyl disulfide, were identified in the fermentation broth. Among these VOCs, methyl thioacetate exhibited multiple nematicidal activities, including contact nematicidal, fumigant, and repellent activities against M. incognita. Methyl thioacetate showed a significant contact nematicidal activity with 87.90% mortality at 0.01 mg/mL by 72 h, fumigant activity in mortality 91.10% at 1 mg/mL by 48 h, and repellent activity at 0.01–10 mg/mL. In addition, methyl thioacetate exhibited 80–100% egg-hatching inhibition on the 7th day over the range of 0.5 mg/mL to 5 mg/mL. These results showed that methyl thioacetate from MCCC 1K02966 control M. incognita with multiple nematicidal modes and can be used as a potential biological control agent.