Viscosity Of Broth Research Articles

Bioethanol production from cassava fed-batch fermentation in pervaporation membrane bioreactor with permeate fractional condensation

A new type pervaporation membrane bioreactor was developed by coupling cassava-based fed-batch fermentation and pervaporation membrane separation technology for bioethanol production. Membrane separation slowed down broth accumulation rate and mitigated ethanol product inhibition. The fermentation time, average sugar consumption rate, ethanol production and ethanol productivity were 131h, 6.6 gL-1h-1, 149.4 gL-1 and 2.5 gL-1h-1, respectively. They were improved by 55.9 %, 73.2 %, 32.3 % and 49.7 %, respectively, compared to those of conventional fed-batch fermentation. The average membrane flux and separation factor were 711.3 gm-2h-1 and 6.5, respectively, with an ethanol concentration of 24.4 wt% in the permeate. Throughout fermentation, the total mass transfer coefficient remained constant at 9.6 × 10-6 ms-1, with the convection mass transfer coefficient decreasing as the broth viscosity increasing while the membrane mass transfer coefficient slightly increasing. A two-stage condensation downstream the membrane achieved fractional vapor recovery. The primary condensate with low ethanol concentration (1.6 wt%) can be used for cassava hydrolyzation, saving 14.0 % of water requirement. Recovering ethanol from the high-concentration (32.6 wt%) secondary condensate required lower energy (46 %) compared to direct recovery from the total permeate.

The effect of viscosity on flavour, mouthfeel and koku enhancement by tastants and yeast extracts in beef broths

This study investigated the effect of viscosity on the dynamic sensory perception of savoury beef broths enriched with savoury enhancers to evaluate the effect of perceived thickness on the koku enhancement effect (i.e. thickness, flavour duration, and complexity). Beef broths were enriched with five combinations of tastants and taste enhancers [none (control); sodium chloride (NaCl); a combination of monosodium glutamate (MSG) and inosine monophosphate (IMP); yeast extract (Kokumax-100); a combination of yeast extract (Kokumax-100) with MSG and IMP]. Three levels of viscosity [low (ƞ=0.72 mPa.s); medium (ƞ=2.94 mPa.s); high (ƞ=12.65 mPa.s)] were obtained by adding different concentrations of locust bean gum (0.00, 0.20, 0.40 w/w%) to beef broths. Temporal-Check-All-That-Apply (TCATA) profiling showed that yeast extract with MSG-IMP enhanced the intensity, richness, body, and duration of savoury sensations of low-viscous beef broths, demonstrating a koku enhancement effect. Increasing the viscosity of broths decreased the perception of taste (i.e. saltiness and savouriness) and flavour (i.e. beef flavour) for most broths, demonstrating a cross-modal suppression of taste and flavour by viscosity. Only broths with yeast extract (Kokumax-100) with MSG-IMP demonstrated stable flavour and taste enhancement across all broth viscosity levels. Koku enhancement by yeast extract (Kokumax-100) with MSG-IMP, MSG-IMP, and NaCl was observed for mouthfeel properties in broths with low and medium viscosity, but not in broths with high viscosity suggesting a suppression of koku enhancement at high broth viscosity. We conclude that (i) yeast extract (Kokumax-100) produces koku enhancement in liquid savoury broths when combined with taste enhancers monosodium glutamate (MSG) and inosine monophosphate (IMP), and (ii) the magnitude of the koku enhancement depends on the presence of taste enhancers and is attenuated at higher broth viscosity.

Investigation of exopolysaccharide formation and its impact on anaerobic succinate production with Vibrio natriegens.

Vibrio natriegens is an emerging host for biotechnology due to its high growth and substrate consumption rates. In industrial processes typically fed-batch processes are applied to obtain high space-time yields. In this study, we established an aerobic glucose-limited fed-batch fermentation with the wild type (wt) of V. natriegens which yielded biomass concentrations of up to 28.4 gX L-1 . However, we observed that the viscosity of the culture broth increased by a factor of 800 at the end of the cultivation due to the formation of 157 ± 20 mg exopolysaccharides (EPS) L-1 . Analysis of the genomic repertoire revealed several genes and gene clusters associated with EPS formation. Deletion of the transcriptional regulator cpsR in V. natriegens wt did not reduce EPS formation, however, it resulted in a constantly low viscosity of the culture broth and altered the carbohydrate content of the EPS. A mutant lacking the cps cluster secreted two-fold less EPS compared to the wt accompanied by an overall low viscosity and a changed EPS composition. When we cultivated the succinate producer V. natriegens Δlldh Δdldh Δpfl Δald Δdns::pycCg (Succ1) under anaerobic conditions on glucose, we also observed an increased viscosity at the end of the cultivation. Deletion of cpsR and the cps cluster in V. natriegens Succ1 reduced the viscosity five- to six-fold which remained at the same level observed at the start of the cultivation. V. natriegens Succ1 ΔcpsR and V. natriegens Succ1 Δcps achieved final succinate concentrations of 51 and 46 g L-1 with a volumetric productivity of 8.5 and 7.7 gSuc L-1 h-1 , respectively. Both strains showed a product yield of about 1.4 molSuc molGlc -1 , which is 27% higher compared with that of V. natriegens Succ1 and corresponds to 81% of the theoretical maximum.

Scale‐related process heterogeneities change properties of high‐cell‐density fermentation broths demonstrated with Escherichia coli B and K‐12 strains

AbstractBACKGROUNDThe scale‐up of Escherichia coli bioprocesses for production of recombinant proteins in the biopharmaceutical industry can affect process performance due to gradients occurring at production scale. The main objective of this study was to investigate the effect of scale‐related process heterogeneities on properties of high‐cell‐density fermentation broths. Two industrially relevant Escherichia coli strains, BL21(DE3) and HMS174(DE3), were used for production of antibody fragments (Fab). To generate these heterogeneities, we used a scale‐down device for cell cultivation, which mimics conditions of production scale (>1000 L) in a 20 L laboratory scale. This setup helps to evaluate the impact of process scale‐up on the resulting fermentation broth properties relevant for primary recovery.RESULTSWe found differences in broth viscosity between the two strains, but also between the standard laboratory‐scale and the scale‐down cultivations used. An increase in cell size was measured in scale‐down cultivations with three orthogonal methods, using scanning electron microscopy, dynamic light scattering and a dispersion analyzer. Strong hints for an altered agglomeration tendency of the cells after scale‐down cultivations were found by measurement of settling velocity and viscoelasticity of the fermentation broth.CONCLUSIONSIn this work, industrially relevant fermentation broths of Escherichia coli cells producing Fab were characterized and represent a relevant basis for rational design of downstream processing operations. The findings show that scale‐related process heterogeneities can alter the phenotype of Escherichia coli cells and thus alter fermentation broth properties relevant for primary recovery. Considering scale‐up effects in upstream processing reflected in downstream processing will help to optimize the entire bioproduction chain in the future. © 2023 The Authors. Journal of Chemical Technology and Biotechnology published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry (SCI).

Efficient production of hyaluronic acid by Streptococcus zooepidemicus using two-stage semi-continuous fermentation

Hyaluronic acid is a kind of mucopolysaccharide that has wide applications in cosmetics, health food, and orthopedics. Using Streptococcus zooepidemicus ATCC 39920 as parent, a beneficial mutant SZ07 was obtained by UV mutagenesis, giving 1.42 g/L hyaluronic acid in shake flasks. To enhance the efficiency of hyaluronic acid production, a semi-continuous fermentation process consisted of two-stage 3–L bioreactors was developed, in which 1.01 g/L/h productivity and 14.60 g/L hyaluronic acid were obtained. To further enhance the titer of hyaluronic acid, recombinant hyaluronidase SzHYal was added into 2nd stage bioreactor at 6 h to reduce the viscosity of broth. The highest hyaluronic acid titer of 29.38 g/L was achieved with a productivity of 1.13 g/L/h at 300 U/L SzHYal after 24 h. This newly developed semi-continuous fermentation process provides a promising strategy for the industrial production of hyaluronic acid and related polysaccharides.

Development of In Situ Product Recovery (ISPR) System Using Amberlite IRA67 for Enhanced Biosynthesis of Hyaluronic Acid by Streptococcus zooepidemicus

High broth viscosity due to the accumulation of hyaluronic acid (HA) causes a limited yield of HA. It is a major problem of HA production using Streptococcus zooepidemicus. Extractive fermentation via in situ product recovery (ISPR) was utilized to enhance the HA production. Resins from Amberlite: IRA400 Cl; IRA900 Cl; IRA410 Cl; IRA402 Cl; and IRA67 were tested for the HA adsorption. IRA67 showed high adsorption capacity on HA. The study of the adsorption via a 2 L stirred tank bioreactor of S. zooepidemicus fermentation was investigated to elucidate the adsorption of HA onto IRA67 in dispersed and integrated internal column systems. The application of a dispersed IRA67 improved the HA production compared to the fermentation without resin addition by 1.37-fold. The HA production was further improved by 1.36-fold with an internal column (3.928 g/L) over that obtained with dispersed IRA67. The cultivation with an internal column shows the highest reduction of viscosity value after the addition of IRA67 resin: from 58.8 to 23.7 (mPa·s), suggesting the most effective ISPR of HA. The improved biosynthesis of HA indicated that an extractive fermentation by ISPR adsorption is effective and may streamline the HA purification.

Cost-Efficient Production of the Sphingan WL Gum by Sphingomonas sp. WG Using Molasses and Sucrose as the Carbon Sources.

The polysaccharide WL gum is produced by the marine microorganism Sphingomonas sp. WG and presents great commercial utility potential in many industries especially in oil industries. However, the high fermentation cost limits its wide application. Therefore, an efficient production system at a lower cost was established using beet molasses to partially replace the commonly used carbon sources. Four different molasses were screened and their composition was investigated. One-factor design and RSM statistical analysis were employed to optimize the WL gum fermentation medium. The effects of molasses on the rheological properties and gene expression of WL gum were also investigated. The results showed that the pretreated beet molasses generated both high broth viscosity and WL gum production (12.94Pa·s and 11.16g/L). Heavy metal ions and ash were found to be the key factors in unpretreated and pretreated molasses affecting WL production. The cost-efficient production medium contained (g/L): sucrose 61.79, molasses 9.95, yeast extract 1.23, K2HPO4 1, MgSO4 0.1, ZnSO4 0.1 and the WL gum production reached 40.25 ± 1.15g/L. The WL gum product WL-molasses showed the higher apparent viscosity, and viscous modulus and elastic modulus than WL-sucrose and WL-mix, which might be related to its highest molecular mass. The higher expressional level of genes such as pgm, ugp, ugd, rmlA, welS, and welG in WL gum synthesis in the mixed carbon source medium causedthe high production and broth viscosity. This work provided a cost-efficient method for WL gum production.

Online measurement of the viscosity in shake flasks enables monitoring of γ-PGA production in depolymerase knockout mutants of Bacillus subtilis with the phosphate-starvation inducible promoter Ppst.

Poly-γ-glutamic acid (γ-PGA) is a biopolymer with a wide range of applications, mainly produced using Bacillus strains. The formation and concomitant secretion of γ-PGA increases the culture broth viscosity, while enzymatic depolymerisation and degradation of γ-PGA decreases the culture broth viscosity. In this study, the recently published ViMOS (Viscosity Monitoring Online System) is applied for optical online measurements of broth viscosity in eight parallel shake flasks. It is shown that the ViMOS is suitable to monitor γ-PGA production and degradation online in shake flasks. This online monitoring enables the detailed analysis of the Ppst promoter and γ-PGA depolymerase knockout mutants in genetically modified Bacillus subtilis 168. The Ppst promoter becomes active under phosphate starvation. The different single depolymerase knockout mutants are ∆ggt, ∆pgdS, ∆cwlO and a triple knockout mutant. An increase in γ-PGA yield in gγ-PGA /gglucose of 190% could be achieved with the triple knockout mutant compared to the Ppst reference strain. The single cwlO knockout also increased γ-PGA production, while the other single knockouts of ggt and pgdS showed no impact. Partial depolymerisation of γ-PGA occurred despite the triple knockout. The online measured data are confirmed with offline measurements. The online viscosity system directly reflects γ-PGA synthesis, γ-PGA depolymerisation, and changes in the molecular weight. Thus, the ViMOS has great potential to rapidly gain detailed and reliable information about new strains and cultivation conditions. The broadened knowledge will facilitate the further optimization of γ-PGA production.

Construction and Performance Evaluation of Dual-Metabolite Oil-Producing Engineering Bacteria Suitable for Low-Permeability Reservoir

Abstract Microbial oil recovery (MEOR) technology has a wide application prospect in the field of enhanced oil recovery in low-permeability reservoirs, and genetic engineering plays an important role. At present, oil-production engineering bacteria constructed by genetic engineering are mainly used to obtain high metabolites and strong environmental adaptability; there are few reports on genetically engineered bacteria with different functional metabolites. However, the bacteria which could efficiently produce one metabolite have a poor ability to produce other metabolites. Therefore, in order to reduce the cost of nutrients and improve the effect of microbial oil recovery, dual-metabolite bacteria have been constructed for low-permeability reservoir. In this paper, four rhamnolipid expression plasmids are extracted from E. coli-produced biosurfactant. Then, the rhamnolipid expression plasmids are transferred into the bacteria-produced biopolymer by electrotransformation, and the metabolite performance of the constructed bacteria is evaluated. Finally, the profile control and displacement performance of the constructed bacteria are studied through low-permeability core experiments. The experimental results show that three bacteria, WS1, WS2, and WS3, are successfully constructed that have preferable ability, among which the bacteria WS2 has the best capacity for producing biopolymer and biosurfactant at the same time. After culturing 96 h, the viscosity of the bacteria WS2 fermentation broth could increase to 42.1 mPa·s, and the surface tension and interfacial tension of the fermentation broth could decrease to 24.3 mN/m and 0.035 mN/m, respectively. This time-varying biological viscosity ensures that the microbial system can enter the low-permeability reservoir at a low injection pressure, and the oil recovery of the low-permeability core could be increased by 10.18% after injecting 0.5 PV WS2 microbial system. The findings of this study can help for better understanding of gene construction and technical support for further popularization and application of MEOR in low-permeability reservoirs.

Identification of the Key Enzymes in WL Gum Biosynthesis and Critical Composition in Viscosity Control.

As an important microbial exopolysaccharide, the sphingan WL gum could be widely used in petroleum, food, and many other fields. However, its lower production is still limiting its wider application. Therefore, to gain insights into the bottlenecks of WL gum production by identifying the key enzymes in the WL gum biosynthesis pathway, more than 20 genes were over-expressed in Sphingomonas sp. WG and their effects on WL gum production and structure were investigated. Compared to the control strain, the WL gum production of welB over-expression strain was increased by 19.0 and 21.0% at 36 and 84 h, respectively. The WL gum production of both atrB and atrD over-expression strains reached 47 g/L, which was approximately 34.5% higher than that of the control strain at 36 h. Therefore, WelB, AtrB, and AtrD may be the key enzymes in WL production. Interestingly, the broth viscosity of most over-expression strains decreased, especially the welJ over-expression strain whose viscosity decreased by 99.3% at 84 h. Polysaccharides’ structural features were investigated to find the critical components in viscosity control. The uronic acid content and total sugar content was affected by only a few genes, therefore, uronic acid and total sugar content may be not the key composition. In comparison, the acetyl degrees were enhanced by over-expression of most genes, which meant that acetyl content may be the critical factor and negatively correlated with the apparent viscosity of WL gum. This work provides useful information on the understanding of the bottlenecks of WL gum biosynthesis and will be helpful for the construction of high WL gum-yielding strains and rheological property controlling in different industries.

Influence of carbon and nitrogen sources on structural features and immunomodulatory activity of exopolysaccharides from Ganoderma lucidum

In this work, the effects of various carbon and nitrogen sources on structural features and immunomodulatory activity of exopolysaccharides (EPS) from Ganoderma lucidum were studied. When glucose and yeast powder were used as carbon and nitrogen sources, the apparent viscosity of fermentation broth and EPS yield and the corresponding sugar content were higher than those cultured with other carbon and nitrogen sources. The main polysaccharide components (20E) with molecular weights ranging from 2.78 × 106 to 3.96 × 106 g/mol were isolated from fermentation broth through 20% ethanol precipitation. Monosaccharide analysis showed that the carbon source significantly affected the monosaccharide compositions of 20E, and the initial carbon source of the medium accounted for the largest proportion of monosaccharide in 20E. The immunoenhancing activities on inducing NF-κB activation by 20E fractions from different carbon and nitrogen combinations exhibited significant differences. Correlation analysis demonstrated that the immunomodulatory activity of 20E fractions was positively correlated with sugar contents and the molar ratios of glucose, while it was negatively correlated with the molar ratios of xylose. As the structural features and biological activities of EPS are highly dependent on the polysaccharide medium composition, this variability can be used advantageously to produce tailor-made polysaccharides with specific applications.

Sustainable production and characterization of medium-molecular weight welan gum produced by a Sphingomonas sp. RW

Welan gum is widely used in food, concrete, and oil recovery for its excellent stability and unique rheological properties. This study describes an engineered Sphingomonas sp. RW strain deficient in the lyase gene (welR) and outlines its application in an industrial by-product fermentation strategy to produce welan gum. The yield of the engineered strain was enhanced to 25.11 ± 0.05 g/L, whereas the broth viscosity (2110 mPas) increased by 281.81%. The welan gum produced by Sphingomonas sp. RW was named medium-molecular weight welan gum (MMWG, ~350 kDa). FT-IR and monosaccharide composition analysis revealed that the MMWG was composed of l-mannose, l-rhamnose, glucuronic acid, and glucose (with mole ratios of 1:4.46:1.26:2.82). Significantly, MMWG had favorable hydroxyl radical scavenging activity and emulsifying capacity. These results provide a strategy for cost-effective welan gum production and enrich the properties of welan gum with different molecular weight.

Hybrid Histidine Kinase WelA of Sphingomonas sp. WG Contributes to WL Gum Biosynthesis and Motility.

Sphingomonas sp. WG produced WL gum with commercial utility potential in many industries. A hybrid sensor histidine kinase/response regulator WelA was identified to regulate the WL gum biosynthesis, and its function was evaluated by gene deletion strategy. The WL gum production and broth viscosity of mutant ΔwelA was only 44% and 0.6% of wild type strain at 72 h. The transcriptomic analysis of differentially expressed genes showed that WelA was mapped to CckA; ChpT, and CtrA in the CckA-ChpT-CtrA pathway was up-regulated. One phosphodiesterase was up-regulated by CtrA, and the intracellular c-di-GMP was decreased. Most genes involved in WL gum biosynthesis pathway was not significantly changed in ΔwelA except the up-regulated atrB and atrD and the down-regulated pmm. Furthermore, the up-regulated regulators ctrA, flaEY, flbD, and flaF may participate in the regulation of flagellar biogenesis and influenced motility. These results suggested that CckA-ChpT-CtrA pathway and c-di-GMP were involved in WL gum biosynthesis regulation. This work provides useful information on the understanding of molecular mechanisms underlying WL gum biosynthesis regulation.

In Situ Corn Fiber Conversion for Ethanol Improvement by the Addition of a Novel Lignocellulolytic Enzyme Cocktail.

Corn mashes have high-viscosity and high-sugar characteristics, which hinders yeast-fermentation efficiency and the ethanol yield increase. The excessive viscosity of corn mash is caused by the unutilized cellulose in corn kernel fiber. A novel lignocellulolytic enzymes cocktail with strong substrate specificity was prepared for high-viscosity, high-sugar corn mash. The in situ conversion of corn mashes with novel lignocellulolytic enzymes at the optimum cellulase dosage of 50 FPU/L resulted in about 12% increased ethanol concentration compared with the reference mash at different batch-fermentation scales. Adding the lignocellulolytic enzymes caused the greatest decrease in viscosity of corn mash and residual sugars by 40.9% and 56.3%, respectively. Simultaneously, the application of lignocellulolytic enzymes increased the value of the dried distiller’s grain with solubles (DDGS) by increasing the protein content by 5.51%. The in situ conversion of cellulose can decrease the fermentation broth viscosity and improve the rheological property, thereby improving the ethanol yield. With the same amount of material, the application of the novel enzymes cocktail can enhance the ethanol yield by more than 12%. A quarter of the ethanol yield increase was due to the further hydrolysis of starch, while three quarters to cellulose. Thus, this technology will increase the net revenue of bioethanol industrialization.

γ-PGA Fermentation by Bacillus subtilis PG-001 with Glucose Feedback Control pH-stat Strategy.

Poly-γ-glutamic acid (γ-PGA) is an important biopolymer with many applications due to its biodegradable and non-toxic characteristics. γ-PGA is produced industrially by fermentation of Bacillus species. The optimal pH range for producing γ-PGA by Bacillus subtilis PG-001 was firstly studied by glucose fed-batch fermentation with non-controlled pH. Result showed that both cell growth and γ-PGA synthesis were repressed when pH was lower than pH 6. Further investigation with γ-PGA fed-batch fermentation showed that pH 6.5 is more suitable for γ-PGA fermentation than pH 7. Under comparable consumption of glutamic acid and glucose, 11.8 g/L γ-PGA and 0.7 g/g yield were achieved by fermentation at pH 6.5, which was significantly higher than 10.5 g/L and 0.56 g/g yield of fermentation at pH 7. In addition, γ-PGA degradation during later phase of fermentation was repressed at pH 6.5 as 9238cP of final broth viscosity was achieved from fermentation at pH 6.5 while it was only 346 cP for fermentation at pH 7. Finally, a glucose feedback control pH-stat strategy was performed for reducing alkali consumption during γ-PGA fermentation, which further increased final γ-PGA concentration to 15.5 g/L with much higher viscosity (11458 cP); meanwhile the consumption of alkali decreased 57%. The fed-batch γ-PGA fermentation with glucose feedback control pH-stat strategy showed high feasibility for industrial scaling-up.

An integrated strategy for recovery and purification of poly-γ-glutamic acid from fermentation broth and its techno-economic analysis

• An integrated strategy for recovery and purification of PGA was developed. • This strategy did not require the use of any exogenous precipitant. • The recovery rate and purity of PGA reached 55% and 94.8%, respectively. • The production cost of $105.58/kg and payback period of 1.55 years were achieved. • Provided an economically feasible strategy for recovery and purification PGA. Poly-γ-glutamic acid (PGA) is a biopolymer with great application prospects and developmental value and which can be produced via microbial fermentation. However, due to the high viscosity of fermentation broth, the recovery and purification of PGA constitute bottlenecks that may limit its use. In this study, an integrated strategy for the recovery and purification of PGA from fermentation broth was developed. Unlike the previously reported ethanol or copper sulfate precipitation strategy, the approach developed in this study includes four unit operations: acidification, plate and frame filtration, cyclic ultrafiltration, and freeze-drying. The strategy does not require the use of any exogenous precipitant, which simplifies the process, lowers costs, and better protects the environment. After optimizing each unit operation, a light yellow powdered PGA was obtained and characterized; its recovery rate, purity, and molecular weight were 55%, 94.8%, and 3,020–5,150 kDa, respectively. Furthermore, the process design and techno-economic analyses were performed at an annual output level of 50 metric tons PGA using SuperPro Designer. A production cost of $105.58/kg and payback period of 1.55 years were achieved. This study provides an economically feasible strategy for the recovery and purification of high-molecular-weight PGA from fermentation broth at an industrial scale.

Effect of biosurfactant sophorolipids on Rhizomucor miehei lipase fermentation by Aspergillus oryzae

In this study, the effect of biosurfactant sophorolipids (SLs) on Rhizomucor miehei lipase (RML) fermentation by Aspergillus oryzae was investigated. With the exogenous addition of 0.3% (w/v) SLs in the initial medium, the RML activity reached 430.0 U/mL, an increase of 25.0% compared to the control group. Subsequently, the physiological metabolic responses of A. oryzae to the addition of SLs were further explored. The results showed that though SLs had almost no effect on the RML secretion, it would affect the morphology of the cells. During the late phase of the fermentation, the proportion of middle pellets, which was generally considered as an energetic and stable state for enzyme production was increased with the addition of SLs. Simultaneously, the viscosity of fermentation broth was reduced, which facilitated the increase of oxygen transfer, thereby improving the RML production. Finally, it could be found that the addition of SLs significantly increased the contents of precursor amino acids, especially for those rank first and second of the RML composition, and it could promote the synthesis of RML.

In-situ corn fiber conversion method unlocks the role of viscosity on enhancing ethanol yield by reducing side-product glycerol

In-situ corn fiber conversion enables to greenly convert corn fiber into bioethanol with saving energy and capital expenditure. However, the remaining challenge is to understand how the additive cellulases improve substrate utilization efficiency but reduce the side-product glycerol concentration. Herein, we investigated a comprehensive responses pattern of cellulases-assisted ethanol production on fermentation properties and transcriptomic level. Two determinants, glucose release rate and viscosity, were identified to govern ethanol/glycerol ratio disturbance. The transcriptomic studies revealed three genes in ethanol synthetic and two genes in synthetic glycerol pathway present remarkably up-regulated (up to 4.28-fold) and down-regulated (up to 4.81-fold) under cellulases condition, respectively. Interestingly, 15 upregulated/downregulated genes were commonly obtained in (non-)cellulases and (non-)sodium-polyacrylate transcriptomic landscape, suggesting the reduced glycerol concentration most likely result from the decreased broth viscosity. This obtained knowledge unmask molecular mechanisms of cellulases-assisted ethanol improvement and empower engineers to rationally optimize the biofuel process for higher ethanol concentration.

Disruption of the Trichoderma reesei gul1 gene stimulates hyphal branching and reduces broth viscosity in cellulase production.

Hyphal morphology is considered to have a close relationship with the production level of secreted proteins by filamentous fungi. In this study, the gul1 gene, which encodes a putative mRNA-binding protein, was disrupted in cellulase-producing fungus Trichoderma reesei. The hyphae of Δgul1 strain produced more lateral branches than the parent strain. Under the condition for cellulase production, disruption of gul1 resulted in smaller mycelial clumps and significantly lower viscosity of fermentation broth. In addition, cellulase production was improved by 22% relative to the parent strain. Transcriptome analysis revealed that a set of genes encoding cell wall remodeling enzymes as well as hydrophobins were differentially expressed in the Δgul1 strain. The results suggest that the regulatory role of gul1 in cell morphogenesis is likely conserved in filamentous fungi. To our knowledge, this is the first report on the engineering of gul1 in an industrially important fungus.

Improving mycelial morphology and adherent growth as well as metabolism of Monascus yellow pigments using nitrate resources

Mycelial adhesion affects cell growth and the production of water-soluble extracellular yellow pigment (EYP) in submerged fermentation with Monascus ruber CGMCC 10910. Two nitrates, NaNO3 and KNO3, were used as nitrogen sources for mitigating mycelial adhesion and improving the production of EYP in this study. The results showed that the adhesion of mycelia in the fermentation broth significantly decreased by adding 5g/L NaNO3, which prevented mycelia from attaching to the inner wall of the Erlenmeyer flask. It was suggested that NaNO3 reduced the total amount of extracellular polysaccharides, increased extracellular proteins, and decreased the viscosity of the fermentation broth. Scanning electron microscopy (SEM) analysis revealed that the mycelial morphology was shorter and more dispersed and vigorous under NaNO3 conditions than under the control conditions. The biomass increased by 49.2% and 45.4% with 5g/L NaNO3 and 6g/L KNO3 treatment, respectively, compared with that of the control, and the maximum production of EYP was 267.1 and 241.8AU350, which increased by 70.0% and 53.9% compared with that of the control, respectively. Simultaneously, the ratios of intracellular yellow pigment to orange pigment increased significantly with 5g/L of NaNO3 addition (p < 0.05). Genetic analysis found that the expression levels of the key genes for Monascus pigment biosynthesis were significantly upregulated by NaNO3 addition (p < 0.05 or p < 0.01). This study provides an effective strategy for the production of water-soluble Monascus yellow pigments.Key Points• Nitrate addition decreased mycelial adhesion and improved cell growth in Monascus pigment fermentation.• The biosynthesis genes of water-soluble extracellular yellow pigment (EYP) were upregulated by nitrate addition.• The mycelial morphology was significantly influenced to enhance EYP biosynthesis with nitrate addition.