Vegan grade medium component screening and concentration optimization for the fermentation of the probiotic strain Lactobacillus paracasei IMC 502® using Design of Experiments.

Lactobacillus paracasei IMC502® is a commercially successful probiotic strain. However, there are no reports that investigate growth medium composition in relation to improved biomass production for this strain. The major outcome of the present study is the design and optimization of a growth medium based on vegan components to be used in the cultivation of Lactobacillus paracasei IMC502®, by using Design of Experiments. Besides comparing different carbon sources, the use of plant-based peptones as nitrogen sources was considered. In particular, the use of guar peptone as the main nitrogen source, in the optimization of fermentation media for the production of probiotics, could replace other plant peptones (e.g. potato, rice, wheat, and soy) which are part of the human diet, thereby avoiding an increase in product and process prices. A model with R2 and adjusted R2 values higher than 95% was obtained. Model accuracy was equal to 94.11%. The vegan-optimized culture medium described in this study increased biomass production by about 65% compared to growth on De Man-Rogosa-Sharpe (MRS) medium. Moreover, this approach showed that most of the salts and trace elements generally present in MRS are not affecting biomass production, thus a simplified medium preparation can be proposed with higher probiotic biomass yield and titer. The possibility to obtain viable lactic acid bacteria at high density from vegetable derived nutrients will be of great interest to specific consumer communities, opening the way to follow this approach with other probiotics of impact for human health.

Xylitol biosynthesis enhancement by Candida tropicalis via medium, process parameter optimization, and co-substrate supplementation.

The present study examines the impact of nitrogen sources (yeast extract, ammonium sulfate peptone, ammonium nitrate, urea, and sodium nitrate), salt solution (0.5 g/L MgSO4, 0.5 g/L KH2PO4, 0.3 g/L CaCl2), trace elements solution (0.1 g/L CuSO4, 0.1 g/L FeSO4, 0.02 g/L MnCl2, 0.02 g/L ZnSO4), operational parameters (temperature, aeration, agitation, initial pH and xylose concentration) and co- substrate supplementation (glucose, fructose, maltose, sucrose, and glycerol) on xylitol biosynthesis by Candida tropicalis ATCC 13803 using synthetic xylose. The significant medium components were identified using the Plackett Burman design followed by central composite designs to obtain the optimal concentration for the critical medium components in shaker flasks. Subsequently, the effect of operational parameters was examined using the One Factor At a Time method, followed by the impact of five co-substrates on xylitol biosynthesis in a 1 L bioreactor. The optimal media components and process parameters are as follows: peptone: 12.68 g/L, yeast extract: 6.62 g/L, salt solution (0.5 g/L MgSO4, 0.5 g/L KH2PO4, and 0.3 g/L CaCl2): 1.23 X (0.62 g/L, 0.62 g/L, and 0.37 g/L respectively), temperature: 30 °C, pH: 6, agitation: 400 rpm, aeration: 1 vvm, and xylose: 50 g/L. Optimization studies resulted in xylitol yield and productivity of 0.71 ± 0.004 g/g and 1.48 ± 0.018 g/L/h, respectively. Glycerol supplementation (2 g/L) further improved xylitol yield (0.83 ± 0.009 g/g) and productivity (1.87 ± 0.020 g/L/h) by 1.66 and 3.12 folds, respectively, higher than the unoptimized conditions thus exhibiting the potential of C. tropicalis ATCC 13803 being used for commercial xylitol production.

Bidirectional fermentation of Monascus and Mulberry leaves enhances GABA and pigment contents: establishment of strategy, studies of bioactivity and mechanistic.

Bidirectional fermentation is a technology that utilizes fungi to ferment medicinal edible substrates, with synergistic and complementary advantages. In this work, a fermentation strategy was established to produce a high yield of γ-aminobutyric acid (GABA) and Monascus pigments (MPs) using Monascus and mulberry leaves (MLs). Firstly, the basic fermentation parameters were determined using single-factor experiments, followed by Plackett-Burman (PB) experimental design to identify MLs, glucose, peptone, and temperature as significant influencing factors. The fermentation parameters were optimized using an artificial neural network (ANN). Finally, the effects of bidirectional fermentation of MLs and Monascus were investigated by bioactivity analysis, microstructure observation, and RT-qPCR. The outcomes showed that the bidirectional fermentation significantly increased the bioactive content and promoted the secondary metabolism of Monascus. The established fermentation conditions were 44.2 g/L of MLs, 57 g/L of glucose, 15 g/L of peptone, 1 g/L of MgSO4, 2 g/L of KH2PO4, 8% (v/v) of inoculum, 180 rpm, initial pH 6, 32 °C and 8 days. The content of GABA reached 13.95 g/L and the color value of MPs reached 408.07 U/mL. This study demonstrated the feasibility of bidirectional fermentation of MLs and Monascus, providing a new idea for the application of MLs and Monascus.

Whey proteins as multifunctional food materials: Recent advancements in hydrolysis, separation, and peptidomimetic approaches.

Whey protein derived bioactives, including α-lactalbumin, ß-lactoglobulin, bovine serum albumin, lactoferrin, transferrin, and proteose-peptones, have exhibited wide ranges of functional, biological and therapeutic properties varying from anticancer, antihypertensive, and antimicrobial effects. In addition, their functional properties involve gelling, emulsifying, and foaming abilities. For these reasons, this review article is framed to understand the relationship existed in between those compound levels and structures with their main functional, biological, and therapeutic properties exhibited either in vitro or in vivo. The impacts of hydrolysis mechanism and separation techniques in enhancing those properties are likewise discussed. Furthermore, special emphasize is given to multifunctional effects of whey derived bioactives and their future trends in ameliorating further food, pharmaceutical, and nutraceutical products. The underlying mechanism effects of those properties are still remained unclear in terms of activity levels, efficacy, and targeted effectiveness. For these reasons, some important models linking to functional properties, thermal properties and cell circumstances are established. Moreover, the coexistence of radical trapping groups, chelating groups, sulfhydryl groups, inhibitory groups, and peptide bonds seemed to be the key elements in triggering those functions and properties. Practical Application: Whey proteins are the byproducts of cheese processing and usually the exploitation of these food waste products has increasingly getting acceptance in many countries, especially European countries. Whey proteins share comparable nutritive values to milk products, particularly on their richness on important proteins that can serve immune protection, structural, and energetic roles. The nutritive profile of whey proteins shows diverse type of bioactive molecules like α-lactalbumin, ß-lactoglobulin, lactoferrin, transferrin, immunoglobulin, and proteose peptones with wide biological importance to the living system, such as in maintaining immunological, neuronal, and signaling roles. The diversification of proteins of whey products prompted scientists to exploit the real mechanisms behind of their biological and therapeutic effects, especially in declining the risk of cancer, tumor, and further complications like diabetes type 2 and hypertension risk effects. For these reasons, profiling these types of proteins using different proteomic and peptidomic approaches helps in determining their biological and therapeutic targets along with their release into gastrointestinal tract conditions and their bioavailabilities into portal circulation, tissue, and organs. The wide applicability of those protein fractions and their derivative bioactive products showed significant impacts in the field of emulsion and double emulsion stabilization by playing roles as emulsifying, surfactant, stabilizing, and foaming agents. Their amphoteric properties helped them to act as excellent encapsulating agents, particularly as vehicle for delivering important vitamins and bioactive compounds. The presence of ferric elements increased their transportation to several metal-ions in the same time increased their scavenging effects to metal-transition and peroxidation of lipids. Their richness with almost essential and nonessential amino acids makes them as selective microbial starters, in addition their richness in sulfhydryl amino acids allowed them to act a cross-linker in conjugating further biomolecules. For instance, conjugating gold-nanoparticles and fluorescent materials in targeting diseases like cancer and tumors in vivo is considered the cutting-edges strategies for these versatile molecules due to their active diffusion across-cell membrane and the presence of specific transporters to these therapeutic molecules.

Transcriptomic profiling reveals differences in the adaptation of two Tetragenococcus halophilus strains to a lupine moromi model medium.

BACKGROUND: Tetragenococcus (T.) halophilus is a common member of the microbial consortia of food fermented under high salt conditions. These comprises salty condiments based on soy or lupine beans, fish sauce, shrimp paste and brined anchovies. Within these fermentations this lactic acid bacterium (LAB) is responsible for the formation of lactic and other short chain acids that contribute to the flavor and lower the pH of the product. In this study, we investigated the transcriptomic profile of the two T. halophilus strains TMW 2.2254 and TMW 2.2256 in a lupine moromi model medium supplied with galactose. To get further insights into which genomic trait is important, we used a setup with two strains. That way we can determine if strain dependent pathways contribute to the overall fitness. These strains differ in the ability to utilize L-arginine, L-aspartate, L-arabinose, D-sorbitol, glycerol, D-lactose or D-melibiose. The lupine moromi model medium is an adapted version of the regular MRS medium supplied with lupine peptone instead of casein peptone and meat extract, to simulate the amino acid availabilities in lupine moromi. RESULTS: The transcriptomic profiles of the T. halophilus strains TMW 2.2254 and TMW 2.2256 in a lupine peptone-based model media supplied with galactose, used as simulation media for a lupine seasoning sauce fermentation, were compared to the determine potentially important traits. Both strains, have a great overlap in their response to the culture conditions but some strain specific features such as the utilization of glycerol, sorbitol and arginine contribute to the overall fitness of the strain TMW 2.2256. Interestingly, although both strains have two non-identical copies of the tagatose-6P pathway and the Leloir pathway increased under the same conditions, TMW 2.2256 prefers the degradation via the tagatose-6P pathway while TMW 2.2254 does not. Furthermore, TMW 2.2256 shows an increase in pathways required for balancing out the intracellular NADH/NADH+ ratios. CONCLUSIONS: Our study reveals for the first time, that both versions of tagatose-6P pathways encoded in both strains are simultaneously active together with the Leloir pathway and contribute to the degradation of galactose. These findings will help to understand the strain dependent features that might be required for a starter strain in lupine moromi.

Isolation, identification, and optimization of the fibrinolytic protease-producing strains.

Cardiovascular complications due to thrombosis have become one of the main causes of death worldwide. The high cost and undesirable side effects of existing thrombolytic agents have led researchers to isolate potential strains that produce fibrinolytic enzymes for therapeutic applications. Fibrinolytic enzymes, especially of microbial origin, are recognized as potential therapeutic candidates for thrombosis. In this study, isolation, identification, and optimization of fibrinolytic protease enzyme-producing strains were performed using fermentative protein sources. Fibrinolytic protease-producing strains were selected by analyzing the isolated strains on skim milk agar medium. The selected strains were examined on blood agar and fibrin plate medium, and the ones showing high enzymatic activity were determined. The strain determined to have the highest activity was identified as Acinetobacter johnsonii TR01 by 16S rRNA analysis. The maximum fibrinolytic protease production of the strain occurred at 60 °C and pH 7.0. Under different medium conditions used for enzyme production, fructose was found to be the best carbon source, while yeast extract and peptone were the best nitrogen sources. It was observed that CaCl2, KH2PO4, and MgSO4 components had a negative effect, while MnCl2 and ZnC4H6O4 components had a positive effect on enzyme production. The medium composition for maximum enzyme activity (8.30 IU/ml) determined by Response Surface Methodology was 14.22 g/L fructose, 11.190 g/L yeast extract, 14.22 g/L peptone, 0.5 g/L MnCl2, and 0.5 g/L ZnC4H6O4.

Modeling and optimization of antibacterial effect of lichen-associated bacteria, Bacillus subtilis KSRLAB3 strain against marine fouling bacteria, Vibrio alginolyticus.

One of the most commonly occurring bacteria, Bacillus subtilis, can produce a wide variety of secondary metabolites. In this study, the antimicrobial effect of B. subtilis KSRLAB3 against Vibrio alginolyticus was optimized using the Plackett-Burman design (PBD) method, response surface methodology (RSM), and genetic algorithm (GA). Initially, the effects of carbon source, nitrogen source, NaCl concentration, pH, temperature, and incubation time on antimicrobial effects were studied. Among the carbon and nitrogen sources investigated, mannose and peptone elicited maximum antimicrobial effect. Using PBD, the most significant variables that influence the antimicrobial effect were identified, including incubation time, peptone concentration, and temperature. The optimum conditions required for attaining maximum antimicrobial effect was identified using the RSM-GA hybrid method, and the optimum condition includes 49.999 h of incubation time, 4.39 g/L of peptone concentration, and 27.629°C of incubation temperature. The confirmatory experiments performed around the optimum condition showed a zone of inhibition of 35 ± 0.52 mm. Methanolic extract also proved the presence of antibacterial lipopeptide surfactin. Therefore, the RSM-GA hybrid method was successfully used in this study to model the antimicrobial effect of B. subtilis KSRLAB3 against V. alginolyticus. The effective inhibition of V. alginolyticus can be investigated further for the development of antifouling coatings.

Evaluation of the Different Nutritional and Environmental Parameters on Microbial Pyrene Degradation by Mangrove Culturable Bacteria.

Mangrove ecosystems play curial roles in providing many ecological services and alleviating global climate change. However, they are in decline globally, mainly threatened by human activities and global warming, and organic pollutants, especially PAHs, are among the crucial reasons. Microbial remediation is a cost-effective and environmentally friendly way of alleviating PAH contamination. Therefore, understanding the effects of environmental and nutritional parameters on the biodegradation of polycyclic aromatic hydrocarbons (PAHs) is significant for the bioremediation of PAH contamination. In the present study, five bacterial strains, designated as Bp1 (Genus Rhodococcus), Sp8 (Genus Nitratireductor), Sp13 (Genus Marinobacter), Sp23 (Genus Pseudonocardia), and Sp24 (Genus Mycolicibacterium), have been isolated from mangrove sediment and their ring hydroxylating dioxygenase (RHD) genes have been successfully amplified. Afterward, their degradation abilities were comprehensively evaluated under normal cultural (monoculture and co-culture) and different nutritional (tryptone, yeast extract, peptone, glucose, sucrose, and NPK fertilizer) and environmental (cetyl trimethyl ammonium bromide (CTAB), sodium dodecyl sulfate (SDS)) parameters, as well with different co-contaminants (phenanthrene and naphthalene) and heavy metals (Cd2+, Cu2+, Fe3+, Ni2+, Mg2+, Mn2+, and Co2+). The results showed that strain Sp24 had the highest pyrene degradation rate (85%) in the monoculture experiment after being cultured for 15 days. Adding nitrogen- and carbon-rich sources, including tryptone, peptone, and yeast extract, generally endorsed pyrene degradation. In contrast, the effects of carbon sources (glucose and sucrose) on pyrene degradation were distinct for different bacterial strains. Furthermore, the addition of NPK fertilizer, SDS, Tween-80, phenanthrene, and naphthalene enhanced the bacterial abilities of pyrene removal significantly (p < 0.05). Heavy metals significantly reduced all bacterial isolates' degradation potentials (p < 0.05). The bacterial consortia containing high bio-surfactant-producing strains showed substantially higher pyrene degradation. Moreover, the consortia of three and five bacterial strains showed more degradation efficiency than those of two bacterial strains. These results provide helpful microbial resources for mangrove ecological remediation and insight into optimized culture strategies for the microbial degradation of PAHs.

Study on the use of bovine blood protein hydrolysate as a peptone in microbial culture media.

Livestock blood is a protein-rich waste byproduct produced during meat production processes in slaughterhouses. Its utilization through conversion into value-added products is an intriguing management strategy. In this study, bovine blood was used to obtain the protein hydrolysate for use as a peptone for microbial growth medium. Lyophilized bovine blood was heat treated to make it susceptible to enzymic hydrolysis, and then enzymatically treated with trypsin (bovine pancreas protease) to produce protein hydrolysate. Physico-chemical features were determined for protein hydrolysate and compared to commercial Merck peptone from meat. Amino acid compositions of bovine blood and commercial peptones were subjected to multivariate analysis based on Euclidean similarity matrix using software PAST. Strains of Staphylococcus aureus 25,923, Pseudomonas aeruginosa 27,853, Staphylococcus aureus 6538 P, Enterococcus faecalis 11,700, Escherichia coli 8739, Klebsiella pneumoniae 13,883, Salmonella typhimurium 14,028 and Listeria monocytogenes 13,932 were used as test microbial strains. Growth of bacteria in culture media based on the peptone from bovine protein hydrolysate was compared to that in corresponding reference media based on commercial peptone. The results of these growth tests were comparable. Growth data were depicted and statistically analyzed using R packages ggplot2 and growthcurver, respectively, providing data fitting a standard form of logistic equation.

Valorization of frying oil waste for biodetergent production using Serratia marcescens N2 and gamma irradiation assisted biorecovery.

BACKGROUND: The complexity, toxicity and abundance of frying oil waste (FOW) render it difficult to be degraded biologically. The aim of the present work was to valorize FOW and investigate the potential use of the produced biosurfactant by Serratia marcescens N2 (Whole Genome sequencing accession ID SPSG00000000) as a biodetergent. RESULTS: Serratia marcescens N2 demonstrated efficient valorization of FOW, using 1% peptone, 20% FOW and 8% inoculum size. Gene annotation showed the presence of serrawettin synthetase indicating that the produced biosurfactant was serrawettin. Zeta potential and Fourier Transform Infrared (FTIR) spectroscopy indicate that the biosurfactant produced was a negatively charged lipopeptide. The biosurfactant reduced the surface tension of water from 72 to 25.7 mN/m; its emulsification index was 90%. The valorization started after 1 h of incubation and reached a maximum of 83.3%. Gamma radiation was used to increase the biosurfactant yield from 9.4 to 19.2 g/L for non-irradiated and 1000 Gy irradiated cultures, respectively. It was noted that the biorecovery took place immediately as opposed to overnight storage required in conventional biosurfactant recovery. Both chemical and functional characteristics of the radiation induced biosurfactant did not change at low doses. The produced biosurfactant was used to wash oil stain; the highest detergency reached was 87% at 60 °C under stirring conditions for 500 Gy gamma assisted biorecovery. Skin irritation tests performed on experimental mice showed no inflammation. CONCLUSION: This study was able to obtain a skin friendly effective biodetergent from low worth FOW using Serratia marcescens N2 with 83% efficient valorization using only peptone in the growth media unlike previous studies using complex media. Gamma radiation was for the first time experimented to assist biosurfactant recovery and doubling the yield without affecting the efficiency.

Nitrogen source as a modulator of the metabolic activity of Pedobacter lusitanus NL19: a transcriptomic approach.

Secondary metabolites (SMs) are compounds with relevant biological activities. Their production under laboratory conditions, especially in broth, is still challenging. An example is the pedopeptins, which are nonribosomal peptides active against some bacteria listed by the WHO for which new antibiotics are urgently needed. Their biosynthesis is inhibited by high concentrations of peptone from casein (PC) in tryptic soy broth (TSB), and we applied a RNA-seq approach to identify Pedobacter lusitanus NL19 cellular pathways modulated by this condition. Results were validated by qPCR and revealed 261 differentially expressed genes (DEGs), 46.3% of them with a predicted biological function. Specifically, high concentration of PC significantly repressed the de novo biosynthesis of biotin (- 60X) and the production of nonribosomal peptide synthetases (NRPS) of pedopeptins (about - 14X), but no effect was observed on the expression of other NRPS. Transcription of a L-Dap synthesis operon that includes a protein with a σ70-like domain was also reduced (about - 7X). High concentrations of PC led to a significant overexpression of MFS and RND efflux pumps and a ferrous iron uptake system, suggesting the redirection of cell machinery to export compounds such as amino acids, sugars and metal divalent cations, alongside with a slight increase of iron import. KEY POINTS: • Higher concentrations of phosphate sources highly repress many operons • High concentrations of peptone from casein (PC) cause biotin's operon repression • High concentrations of PC downregulate the production of peptides of unknown function.

An Evaluation of Different Types of Peptone as Partial Substitutes for Animal-derived Serum in Vero Cell Culture.

Vero cells are one of the most frequently used cell types in virology. They can be used not only as a vehicle for the replication of viruses, but also as a model for investigating viral infectivity, cytopathology and vaccine production. There is increasing awareness of the need to limit the use of animal-derived components in cell culture media for a number of reasons, which include reducing the risk of contamination and decreasing costs related to the downstream processing of commercial products obtained via cell culture. The current study evaluates the use of protein hydrolysates (PHLs), also known as peptones, as partial substitutes for fetal bovine serum (FBS) in Vero cell culture. Eleven plant-based, two yeast-based, and three casein-based peptones were assessed, with different batches evaluated in the study. We tested the effects of three concentration ratios of FBS and peptone on Vero cell proliferation, four days after the initial cell seeding. Some of the tested peptones, when in combination with a minimal 1% level of FBS, supported cell proliferation rates equivalent to those achieved with 10% FBS. Collectively, our findings showed that plant-based peptones could represent promising options for the successful formulation of serum-reduced cell culture media for vaccine production. This is especially relevant in the context of the current COVID-19 pandemic, in view of the urgent need for SARS-CoV-2 virus production for certain types of vaccine. The current study contributes to the Three Rs principle of reduction, as well as addressing animal ethics concerns associated with FBS, by repurposing PHLs for use in cell culture.

Optimization of culture conditions for enhanced Dunaliella salina productions in mixotrophic culture.

Dunaliella salina (D. salina) is a green microalga known for its tendency to produce lipid and ß-carotene. Fatty acid profile, lipid and ß-carotene productions of the microalga D. salina cultivated under different mixotrophic conditions were assayed. Notably, in spite of a broad spectrum of substrates served, mixotrophic cultivations slightly affected the fatty acid composition, and as a result C16:0 and C18:0, C18:1, C18:2 and C18:3 were identified as main fatty acids. Lipid in dry weight biomass (DWB) hit a high of 24.3% at 5% of NaCL and linolenic acid in lipid reached a peak of 9.15% at 15% of NaCL in medium containing glucose and equal amounts of yeast extract and soy bean powder. One-factor-at-a-time was applied to elucidate the substrates which had noticeable impacts on ß-carotene production. Glucose, meat peptone, titanium dioxide nanoparticles (TiO2 NPs), pH 7.5 and 5% NaCL were identified as key process parameters impacting ß-carotene production. Following, the concentration of glucose, meat peptone and TiO2 NPs were optimized by using response surface method. The highest content of ß-carotene, 25.23 mg/g DWB, was obtained in medium composed of (g/L); 22.92 glucose, 5 meat peptone and 0.002 TiO2 NPs.

Comparative transcriptome analysis for the biosynthesis of antioxidant exopolysaccharide in Streptococcus thermophilus CS6.

BACKGROUND: Food grade Streptococcus thermophilus produces biological exopolysaccharides (EPSs) with great potential with respect to catering for higher health-promoting demands; however, how S. thermophilus regulates the biosynthesis of EPS is not completely understood, decelerating the application of these polymers. In our previous study, maltose, soy peptone and initial pH were three key factors of enhancing EPS yield in S. thermophilus CS6. Therefore, we aimed to investigate the regulating mechanisms of EPS biosynthesis in S. thermophilus CS6 via the method of comparative transcriptome and differential carbohydrate metabolism. RESULTS: Soy peptone addition (58.6 g L-1 ) and a moderate pH (6.5) contributed to a high bacterial biomass and a high EPS yield (407 mg L-1 ). Maltose, soy peptone and initial pH greatly influenced lactose utilization in CS6. Soy peptone addition induced a high accumulation of mannose and arabinose in intracellular CS6, differential monosaccharide composition (mannose, glucose and arabinose) in EPS and high radical [2,2-diphenyl-1-picrylhydrazyl, superoxide and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)] scavenging activities. Carbohydrate transportation, sugar activation and eps cluster-associated genes were differentially expressed to regulate EPS biosynthesis. Correlation analysis indicated high production of EPSs depended on high expression of lacS, galPMKUTE, pgm, gt2-5&4-1 and epsLM. CONCLUSION: The production of antioxidant EPS in S. thermophilus CS6 depended on the regulation of galactose metabolism cluster and eps cluster. The present study recommends a new approach for enhancing EPS production by transcriptomic regulation for further food and health application of EPS. © 2022 Society of Chemical Industry.

Enhancement production of lipid and unsaturation of fatty acids in Cryptococcus humicola via addition of calcium ion.

Lipids synthesized by oleaginous yeasts are considered to be the best candidates for biodiesel production. Cryptococcus humicola as an oleaginous yeast accumulated lipid in cells. In order to optimize the conditions for lipid production, different carbon and nitrogen sources were used and metals were added into the medium. Ca2+ addition increased the lipid production greatly. Xylose and peptone were optimal carbon source and nitrogen source, respectively for lipid accumulation. Response surface experiment results revealed that the accumulation of lipid could be maximized when the xylose, peptone and Ca2+ concentration was 61 g/L, 4.31 g/L, 0.67 mmol/L. C16 and C18 fatty acid account for about 91% of the total fatty acids. The most abundant fatty acid was oleic acid (42.68%), followed by palmitic acid (29.7%) and stearic acid (13.87%). The addition of Ca2+ increased the content of unsaturated fatty acids (such as C16:1 and C18:1) and improved the unsaturation of fatty acids. Quantitative real time PCR analysis revealed that expression of genes related to lipid biosynthesis showed up-regulated by Ca2+ treatment. This study provided a strategy for increase in lipid production and content of unsaturated fatty acids.

Effect of culture medium optimization on the secondary metabolites activity of Lysobacter antibioticus 13-6.

Fermentation products of Lysobacter antibioticus 13-6 have antagonistic activity against devastating phytopathogenic bacerium Xanthomonas oryzae pv. oryzicola. The production of Lysobacter antibioticus 13-6 secondary metabolites was increased by optimizing the fermentation medium; using a single-factor screening test, Plackett-Burman Design, and Box-Behnken Design. The medium's final formulation for active secondary metabolites high-yield included peptone 5 g/L, glucose 4.73 g/L, MgSO4·7H2O 2.33 g/L, and K2HPO4 2.21 g/L. We compared phenazine-1-carboxylic acid (PCA) contents of L. antibioticus 13-6 in the initial and optimized mediums through HPLC. It was found PCA contents of the optimized medium are two folds more than in the initial medium. We also detected the relative expression of five phenazine genes of L. antibioticus 13-6 via RT-qPCR, and it was found that genes: phzB, C, S, and NO1 have more significant expression compared with the initial medium, while gene phzD has found just significant. Further, we revealed that the optimal fermentation conditions for secondary metabolites were: fermentation time 60 hours, shaking speed 160 rpm, inoculum size 3%, and the initial pH = 7.0. In the end, it was determined that the antimicrobial activity and quality of L. antibioticus 13-6 secondary metabolites were increased by about 41.75% and 2-times, respectively, after the optimization of the fermentation medium.

The effect of carbon, nitrogen and iron ions on mono-rhamnolipid production and rhamnolipid synthesis gene expression by Pseudomonas aeruginosa ATCC 15442.

Pseudomonas spp. are the main producers of rhamnolipids. These products have applications in pharmaceuticals, cosmetics, food industry and bioremediation. The biosynthesis of rhamnolipids is influenced by nutrient composition, pH and temperature. In this study, the impact of nutrients on the expression levels of rhamnolipid synthesis genes was evaluated in P. aeruginosa ATCC 15442. Glucose and glycerol were used as carbon sources; while, NaNO3, NH4NO3 and yeast extract/peptone were employed as nitrogen sources. The effect of different concentrations of Fe2+ and Fe3+ on rhamnolipid synthesis genes was also evaluated. Highest biosurfactant production was obtained in minimal medium supplemented with glucose, NaNO3 and Fe2+. Two rhamnolipid synthesis genes, rhlA and rhlB, were amplified with PCR. CapLC ESI-Ion trap-MS/MS detected only mono-rhamnolipid Rha-C10-C10 in the extract. Although similar induction levels were recorded in the presence of 0.05 g/L iron ions, the presence of Fe2+ resulted in higher expression levels than Fe3+ at concentrations equivalent to 0.025 and 0.075 g/L.

Direct transesterification of fatty acids produced by Fusarium solani for biodiesel production: effect of carbon and nitrogen on lipid accumulation in the fungal biomass.

AIMS: The present work aims to explore a new oleaginous Fusarium isolate potential to accumulate lipids in its biomass from inexpensive substrates. In addition, impacts of carbon and nitrogen sources and their ratios on lipid production by the interested fungal isolate were also studied. METHODS AND RESULTS: Lipid was assayed by sulfo-phospho-vanillin colorimetric method. Among 11 Fusarium isolates obtained on potato dextrose agar from rhizosphereic soils, Fusarium RAS18 was selected as the highest producer that accumulates above 20% lipid. It was identified based on phenotypic characterization and the internal transcribed spacer sequence as Fusarium solani, that was recorded in the GenBank database under the accession number MK167372.1. The optimized lipid yield (34·5%) is obtained using glycerol (35 g l-1 ) and peptone (1·5 g l-1 ) as carbon and nitrogen sources respectively. The produced fatty acid methyl esters (biodiesel) is composed of linoleic acid (56·81%), palmitic acid (17·81%), oleic acid (11·81%) and stearic acid (11·12). The unsaturated fatty acids accounted for 69% and this is nearly similar to the plant oils commonly used in biodiesel production. CONCLUSIONS: These findings suggest the applicability of F. solani RAS18 as a promising strain to accumulate lipids from glycerol as a feedstock for biodiesel production. SIGNIFICANCE AND IMPACT OF THE STUDY: Fusarium solani RAS18 is a new oleaginous fungal isolate that is able to produce lipid (34·5%, g g-1 ) from glycerol. Glycerol is a cheap substrate and is formed as a byproduct from transesterification process and others industries. Thus, recyclation of glycerol for lipid production by micro-organisms is an important point of economic view. Direct transesterification of the produced fatty acids indicated its similarity to the plant oil composition used in biodiesel production. So, F. solani RAS18 might be a potential lipid source as a feedstock for biodiesel production.

Consistent production of kojic acid from Aspergillus sojae SSC-3 isolated from rice husk.

A consistent kojic acid producing fungal strain has been isolated from rice husk using glucose-peptone medium. The isolate was identified as Aspergillus sojae SSC-3 on 18S rDNA analysis. A. sojae was capable of producing substantially good amount of kojic acid, however the production was varying from batch to batch. In order to obtain consistent, repeated and high levels of kojic acid, monospore isolation procedures was adopted. The highest production of kojic acid obtained was 12 ± 2 g/L in 120 h with sucrose (10%) and yeast extract (0.5%) as carbon and nitrogen source respectively. The process was scale up to 10 L fermenter size which repeatedly resulted in the production of 18 ± 2 g/L of kojic acid in 96 h. Kojic acid was recovered (> 82%) from the fermentation broth with > 99% purity. Best to our knowledge this is the first report were kojic acid production is reported from Aspergillus sojae strain.

Five-factor-at-a-time (FFAT) approach for optimal production of an extracellular RNase from Bacillus safensis RB-5.

A Gram-positive, rod-shaped, endospore-forming, and RNA-degrading bacterium RB-5 was isolated from a soil sample. Based on 16-rDNA gene sequence, the bacterium RB-5 was identified as Bacillus safensis (Accession number KX443714.1). The bacterium appeared to be related to Bacillus safensis KL-052, an other-member of genus Bacillus. One-factor-at-a-time (OFAT) and Response Surface Methodology (RSM) statistical approaches were used to optimize the fermentation broth to obtain an improved extracellular RNase production from B. safensis RB-5. These approaches improved RNase activity of B. safensis KL-052 from 4.26 to 7.85 U/mL. The OFAT approach was used to study the effects of supplementation of carbon, nitrogen and physical conditions, which included temperature, pH and agitation rate on extracellular RNase production by B. safensis KL-052. Five variables screened by Central Composite Design (CCD) were employed to evaluate their interactive effects on RNase production by the organism. CCD selected 25 factorial values obtained by the statistical approach were peptone 1.13% (w/v), sodium nitrate 1.13% (w/v), MgSO4 0.06% (w/v), pH 8.5, and temperature 35 °C for RNase production by B. safensis. The highest predicted value of RNase was 7.05 U/ml while actual obtained value was 7.85 U/ml that was ∼84% and 1.84-fold higher than OFAT approach.