Fermentation Conditions Research Articles

Engineered production of 5-aminolevulinic acid in recombinant Escherichia coli BL21.

5-aminolevulinic acid (ALA) is a non-protein amino acid that has been widely used in the fields of medicine and agriculture. This study aims to engineer the C5 pathway of the ALA biosynthesis in Escherichia coli BL21 to enhance ALA production. The ALA synthase genes gltX, hemA, and hemL were overexpressed in E. coli BL21 to lead to the increase in the production of ALA. The sRNA RyhB was also overexpressed to downregulate the expression of ALA dehydratase to reduce the downstream bioconversion of ALA to porphobilinogen. Next, the gene arcA was knocked out by CRISPR-Cas9 technology to open the TCA cycle to promote the respiratory metabolism of the strain to reduce the feedback inhibition of heme to ALA. The fermentation conditions of the engineered strain were optimized by response surface experiments. The time-course analysis of the ALA production was carried out in a 1 L shake flask. Through these efforts, the production of ALA in engineered strain reached 2953 mg/L in a 1 L shake flask. This study contributes to the industrial production of ALA by the engineered E. coli in the future.

Investigation of the Fermentation Process of Moringa oleifera Leaves and Its Effects on the Growth Performance, Antioxidant Capacity, and Intestinal Microbiome of Procambarus clarkii

Moringa oleifera is renowned for its high antioxidant activity. However, few studies have been conducted on its effects on aquatic animals. The aim of this experiment was to investigate the optimal fermentation process of M. oleifera leaves and to evaluate the effects of fermented M. oleifera leaves on crayfish (9.11 ± 0.3 g) in terms of growth performance, antioxidant capacity, and gut microbiological parameters. By optimizing the fermenting material/water ratio, fermentation time, temperature, and strain, the optimal fermentation conditions of a 10% water ratio + 48 h + 30 °C + inoculation with 2% B. amyloliquefaciens (107 CFU mL−1) were obtained. These conditions resulted in notable increases in the contents of the total protein, total phenols, flavonoids, and amino acids (p < 0.05) while also leading to a notable decrease in the content of tannins in contrast to those of unfermented M. oleifera leaves (p < 0.05). The fermented M. oleifera (FMO) leaves were incorporated at five concentrations, including 0% (control (CT)), 0.25% (0.25FMO), 0.5% (0.5FMO), 1% (1FMO), and 2% (2FMO). The results showed that the 1FMO group performed better in terms of the final body weight (FBW), weight gain rate (WGR), and specific weight gain rate (SGR) compared with the CT group (p < 0.05). In addition, amylase and lipase activities were significantly higher in the 1FMO and 2FMO groups compared with the other groups (p < 0.05). The fermented M. oleifera leaves significantly increased the catalase (CAT) activity in the crayfish (p < 0.05). The superoxide dismutase (SOD) activity was significantly increased in the 0.25FMO, 1FMO, and 2FMO groups, and the malondialdehyde (MDA) content was significantly decreased while the glutathione peroxidase (GSH-PX) content was significantly increased in the 0.5FMO, 1FMO, and 2FMO groups (p < 0.05). Furthermore, the 1FMO group was observed to significantly increase the abundance of Firmicutes while simultaneously reducing the abundance of Aeromonas (p < 0.05) and adjusting the structure of the intestinal microbiome. In conclusion, this study established the optimal fermentation conditions for M. oleifera and obtained a product with high nutrient and low tannin contents. Furthermore, the incorporation of 1% FMO was demonstrated to facilitate growth, enhance the antioxidant capacity, and optimize the gut microbiology in crayfish.

Exploring the Acetobacteraceae family isolated from kombucha SCOBYs worldwide and comparing yield and characteristics of biocellulose under various fermentation conditions

Bacterial cellulose (BC) is a cellulosic biopolymer produced by specific acetic acid bacteria during kombucha fermentation. In this study, bacterial cellulose-producing strains were isolated from four different global kombucha SCOBY samples obtained from markets in the Netherlands, America, China, and Iran. The strains were identified using biochemical and molecular techniques. The ability of species to produce BC was evaluated under both static and stirred fermentation conditions. Seven dominant strains from the Acetobacteraceae family and the genus of Komagataeibacter and Gluconacetobacter were identified and submitted to NCBI gene bank archives: K. xylinus CH1, K. sucrofermentans IR2, K. intermedius IR3, K. cocois AM2, K. sucrofermentans NE4, K. cocois NE6, and G. liquefaciens NE7. Among these, K. intermedius IR3, isolated from local Iranian SCOBY, exhibited the highest BC production yield at 5.733 ± 0.170 gL−1 under static fermentation conditions. On the other hand, K. xylinus CH1, from Chinese SCOBY, had the highest yield under stirred conditions, producing 12.689 ± 0.808 gL−1 of BC. The BC production yield of both K. xylinus CH1 and K. intermedius IR3 under stirred conditions was 3 and 1.3 times more than static conditions, respectively. Despite the yield differences, static fermentation demonstrated superior physicochemical characteristics; such as moisture content, water holding capacity, and crystallinity degree, compared to stirred. Therefore, depending on the intended application in industry and specific criteria, both products could serve as functional substitutes in food and medicine sectors.

From Waste to Protein: Optimizing Rice Husk Utilization for Sustainable Single Cell Protein Production

The increasing global demand for protein has prompted a search for sustainable and cost-effective alternatives to traditional animal and dairy sources. Single-cell protein (SCP) produced from microorganism’s offers a promising solution due to its high protein content and rapid growth rates. This study addresses the challenge of utilizing rice husk, an abundant yet underutilized agricultural waste in Nigeria, as a substrate for SCP production. Through optimization of the production of SCP using Bacillus sp. AT-b3 and Bacillus sp. CMF 12 as model organisms. Molecular identification through 16S rRNA sequencing confirmed Bacillus sp. AT-b3 and Bacillus sp. CMF 12 as the most potent isolates for SCP production. Optimization studies were conducted using the One-Factor-at-a-Time (OFAT) method to determine the ideal fermentation conditions. The results showed that the optimal temperature for SCP production was 40 °C, with Bacillus sp. AT-b3 demonstrating superior production efficiency. pH optimization revealed that neutral pH (pH 7) was ideal for maximizing SCP production, with Bacillus spp. AT-b3 outperforming Bacillus sp. CMF 12 at this pH. Substrate concentration studies indicated that 2.0% was optimal for SCP production, and incubation time optimization indicated 48 hours as the optimal period for maximum yield. Amino acid profiling of the SCP produced showed significant variations between the two isolates. Bacillus sp. CMF 12 was richer in essential amino acids like Arginine and Methionine, while Bacillus sp. AT-b3 had a higher Glycine content at (p<0.05). The findings of this study suggest that both strains have potential applications in nutritional supplements, with Bacillus sp. AT-b3 being particularly suited for industrial-scale SCP production. This study concludes that Bacillus sp. AT-b3 is an efficient SCP producer under optimal conditions of neutral pH, moderate temperature, and appropriate substrate concentration. Further research is recommended to explore pilot-scale production, alternative substrates, and comprehensive safety assessments.

Saga of Soggy Sauerkraut

The creation of undesirable (soggy) sauerkraut resulted in the loss of $1,000,000 worth of organic sauerkraut in 2022, which prompted a multistep investigation of the cause and potential solution. The cause of this condition has been previously reported as unique fermentation conditions and the lack of key trace nutrients essential for cabbage (Brassica oleracea var. capitata) cell wall integrity. Because the condition was limited to organic sauerkraut in 2022, this investigation initially focused on differences in fermentation conditions between organic and conventional sauerkraut. No differences in fermentation conditions accounted for the condition; therefore, attention was focused on analyzing the mineral content of cabbage grown for sauerkraut production that pinpointed a deficiency in critical micronutrients such as iron, copper, manganese, boron, and zinc. This deficiency was traced to the use of poultry manure that was contaminated with glyphosate residue from conventionally fed turkeys and chickens that consumed genetically engineered (GE) feed and used as the fertilizer for organic cabbage production. The presence of glyphosate, a potent mineral chelator and antibiotic, was identified as a significant factor that impairs the absorption and physiological function of essential minerals in the shikimate metabolic pathway whereby cell walls and lignin are produced, thus compromising the structural quality of the sauerkraut. After this discovery, the study progressed to evaluate various remediation strategies aimed at eliminating glyphosate from the soil and restoring nutrient uptake. Corn grain and silage were selected as the test crops for this phase. Among the tested remediation solutions were raw sauerkraut juice containing Lactobacillus plantarum, which is reported to degrade glyphosate in the rumen of dairy cows and two patented proprietary microbial mixtures, PB027 and PB027SK, that degrade glyphosate by all three of the known metabolic pathways. These treatments were specifically formulated to degrade residual glyphosate in the soil. The results showed that these interventions could reduce soil glyphosate levels by 80% to 90% within 6 to 7 months to significantly enhance both the yield and quality of corn grain and silage. The increase in corn grain yield from glyphosate degradation on the Shiocton silt loam soil was 907.89 kg·ha−1 (13.5 bushels/acre). The increase in yield on the irrigated Kidder sandy loam soil was quantified at 726.31 kg·ha−1 (10.8 bushels/acre) for corn grain and 6.62 t·ha−1 (2.68 t/acre) for silage, with an additional improvement in silage feed quality beneficial for milk production. The findings underscore the importance of addressing both micronutrient sufficiency and glyphosate residue in soil to ensure the optimal growth of cabbage and the quality of sauerkraut produced. By successfully identifying manure as a subtle source of nutrient immobilization and implementing effective soil remediation techniques, this research highlights a clear path forward for improving crop yield and quality to ultimately enhance the structural integrity and consumer acceptance of sauerkraut. This study has broader applications for the nutritional content and crop yields of many organic crops that use conventional poultry and animal manures that may contain glyphosate in desiccated plant tissues or GE feeding operations.

Controlling strategies of methanol generation in fermented fruit wine: Pathways, advances, and applications.

Methanol is widely existed in fermented fruit wines (FFWs), and the concentration is excessive at times due to inappropriate fermentation conditions. Methanol is neurotoxic, and its metabolites of formaldehyde and formic acid can cause organic lesions and central respiratory system disorders. FFWs with unspecified methanol limits are often produced with reference to grape wine standards (250/400mg/L). To clarify the causes of methanol production in FFWs and minimize the methanol content, this study summarizes the current process methods commonly applied for methanol reduction in FFWs and proposes novel potential controlling strategies from the perspective of raw materials (pectin, pectinase, and yeast), which are mainly the low esterification modification and removal of pectin, passivation of the pectinase activity, and the gene editing of yeast to target the secretion of pectinases and modulation of the glycine metabolic pathway. The modified raw materials combined with optimized fermentation processes will hopefully be able to improve the current situation of high methanol content in FFWs. Methanol detection technologies have been outlined and combined with machine learning that will potentially guide the production of low-methanol FFWs and the setting of methanol limits for specific FFW.

DMSO suppresses duclauxin biosynthetic pathway in Talaromyces sp. (strain IQ-313) and untaps terpenoids, polyketides and meroterpenoids biosynthesis

Talaromyces sp. (strain IQ-313) produces duclauxin-type molecules under standard fermentation conditions in rice and oat cereal. Such molecules are of great interest due to their structural complexity and biological activity. Interestingly, application of an OSMAC (One Strain Many Compounds) approach, revealed that the biosynthetic machinery of this fungus can be directed towards the production of other types of molecules, while completely suppressing the biosynthesis of duclauxin (1). To exploit the metabolic potential of Talaromyces sp. (strain IQ-313), it was grown on solid media supplemented with dimethyl sulfoxide (DMSO), an ectopic epigenetic modulator. Metabolomic analysis of the fermentation extract from the strain grown under standard and DMSO-supplemented conditions, using molecular networking, revealed notable differences in the profiles. Chemical investigation of the extract obtained under abiotic stress led to the isolation and characterization of 15 molecules not detected under standard conditions, including nine polyketides, one sesquiterpenoid, two sesterterpenoids, and three meroterpenoids. Among these, talaromophylane (2) an eremophilane sesquiterpenoid, and talaroisochromane (8), an oxoisochromen, have not been previously reported in the literature. The structures of all isolates were established using a combination of spectroscopic and spectrometric data. The absolute configuration of compound 1 was established based on the analysis of NOESY interactions and by comparison of the experimental and theoretical electronic circular dichroism (ECD) curves.

Screening and Selection of a New Medium and Culture Conditions for Diosgenin Production via Microbial Biocatalysis of SYt1

Diosgenin (DSG) is a phytosterol saponin mainly found in Dioscorea zingiberensis C.H. Wright. It has shown promising results in treating various diseases such as cancer, diabetes, arthritis, asthma, and cardiovascular diseases. Diosgenin is also an important medicinal chemical for synthesizing various steroid medicines. The production of diosgenin by acid hydrolysis generates a large amount of wastewater, leading to severe environmental pollution. However, producing diosgenin through microbial fermentation can effectively reduce environmental pollution. Numerous studies have demonstrated that various microorganisms can produce diosgenin via solid-state fermentation. Nevertheless, due to the complexity, high maintenance costs, uneven heat production, and other characteristics of solid-state fermentation, it is not commonly used in the industrial production of diosgenin. In contrast, liquid fermentation offers advantages such as simple operation, easy maintenance, and stable fermentation, making it more suitable for the industrial production of diosgenin. However, few studies have focused on producing diosgenin using liquid fermentation. In this study, endophytic Bacillus licheniformis SYt1 was used to produce diosgenin via liquid fermentation, with Dioscorea tuber powder as a substrate. Soxhlet extraction and silica gel column chromatography were employed to identify the diosgenin from the liquid fermentation products. Suitable fermentation conditions were screened and identified. The environmental variables that significantly affect the diosgenin yield were determined by the Plackett–Burman design (P-BD) with eight factors. The three factors (peptone, yeast extract powder and inorganic salt) with the greatest influence on the diosgenin yield were selected and further optimized using a response surface methodology (RSM). The final culture conditions were determined to be 35.79 g/L of peptone, 14.56 g/L of yeast extract powder, and 1.44 g/L of inorganic salt. The yield of diosgenin under these conditions was 132.57 mg/L, which was 1.8 times greater than the yield under pre-optimization conditions. This effective, clean, and promising liquid fermentation method possesses the potential to replace the traditional acid hydrolysis method for the industrial production of diosgenin.

Effect of Mixed Strains on Microbial Community and Flavor Metabolites in Fermentation Process of Chi-Flavor Baijiu

The distinct flavor of chi-flavor baijiu (CFB) has garnered significant attention in China. After the optimization of fermentation conditions, Pichia anomala and Lactobacillus plantarum were introduced into the fermentation process to enhance the flavor. Samples inoculated with these mixed strains (SY) exhibited higher levels of alcohol (from 33.04 to 178.55 mg/L) and esters (from 49.51 to 130.20 mg/L) compared to the control group (KB). In SY, P. anomala and L. plantarum were the predominant microorganisms, while Pediococcus and Saccharomyces were more prevalent in KB. Moreover, 68 volatile flavor compounds were detected in SY, as opposed to 64 in KB. Notably, Pichia showed a positive correlation with key flavor compounds. The synergistic fermentation with exogenous strains led to a 52.38% increase in phenethyl alcohol and a 4.91% increase in ethyl lactate. Additionally, the levels of other flavor compounds, like ethyl acetate, γ-nonanolactone, and (E)-2-octenal, also increased. The results demonstrated that the addition of P. anomala and L. plantarum to the fermentation process of CFB significantly increased the contents of flavor compounds. This research reveals valuable insights into flavor enhancement and the microbial community dynamics in CFB production.

Degradation of Anti-Nutritional Factors in Maize Gluten Feed by Fermentation with Bacillus subtilis: A Focused Study on Optimizing Fermentation Conditions

Degradation of Anti-Nutritional Factors in Maize Gluten Feed by Fermentation with Bacillus subtilis: A Focused Study on Optimizing Fermentation Conditions

Optimization of fermentation conditions for physcion production of Aspergillus chevalieri BYST01 by response surface methodology.

This study aimed to optimize the culture conditions of the termite-derived fungus Aspergillus chevalieri BYST01 for the production of physcion, a characteristic component of the traditional herb rhubarb, which has been commercially approved as a botanical fungicide in China. First, potato dextrose broth was screened as the suitable basal medium for further optimization, with an initial yield of 28.0 mg/L. Then, the suitable carbon source, fermentation time, temperature, pH value, and the rotary shaker speed for physcion production were determined using the one-variable-at-a-time method. Based on the results of single factors experiments, the variables with statistically significant effects on physcion production were further confirmed using the Plackett-Burman design (PBD). Among the five variables, temperature, initial pH, and rotary shaker speed were identified as significant factors (P < 0.05) for physcion productivity in the PDB and were further analyzed by response surface methodology (RSM). Finally, we found that the maximum physcion production (82.0 mg/L) was achieved under the following optimized conditions:initial pH 6.6, rotary shaker speed of 177 rpm, temperature of 28 °C, and glucose concentration of 30 g/L in PDB medium after 11 d of fermentation. The yield of physcion under the optimized culture conditions was approximately threefold higher than that obtained using the basal culture medium. Furthermore, the optimum fermentation conditions in the 5-L bioreactor achieved a maximal physcion yield of 85.2 mg/L within 8 d of fermentation. Hence, response surface methodology proved to be a powerful tool for optimizing physcion production by A. chevalieri BYST01. This study may be helpful in promoting the application of physcion produced by A. chevalieri BYST01 to manage plant diseases.

Preparation of safflower fermentation solution and study on its biological activity

IntroductionSafflower, a traditional Chinese medicine, is rich in chemical components including flavonoids, polysaccharides, and alkaloids. It exhibits pharmacological effects such as antioxidant, anti-inflammatory, anti-tumor, and anti-thrombosis properties, making it a valuable resource in the medical field. Furthermore, due to its antioxidant and anti-inflammatory effects, safflower is increasingly being utilized in the cosmetics industry.MethodsIn this study, yeast was employed to ferment safflower, and the optimal fermentation conditions were established through single-factor experiments and response surface methodology. Subsequently, the antioxidant and anti-inflammatory efficacy of the safflower fermentation solution was assessed using both cellular and zebrafish models. Finally, the safety of the safflower fermentation solution was evaluated through a cosmetic eye irritation test.ResultsFrom a total of 20 yeast strains, YF-5 was identified as the dominant strain for safflower fermentation. By optimizing the fermentation conditions, it was established that the optimal parameters for YF-5 fermentation of safflower are as follows: a fermentation temperature of 36.55°C, a material-to-liquid ratio of 1:20.46, a fructose concentration of 6.20%, a fermentation duration of 72 h, and an inoculum volume of 4%. The biological activities of safflower, including its antioxidant and anti-inflammatory properties, were enhanced through yeast fermentation. In HaCaT cell and zebrafish oxidative damage assays, safflower fermentation solution inhibits the production of malondialdehyde (MDA) and increases superoxide dismutase (SOD) activity as well as total antioxidant capacity (T-AOC). In the RAW264.7 cell inflammatory damage assays, a 20% safflower fermentation solution was found to inhibit the release of TNF-α and NO in the inflammatory model, with inhibition rates of 30.94 and 28.86%, respectively. In the zebrafish inflammatory damage assays, the quantity of fluorescent neutral proteins in the 5% safflower fermentation solution was 0.7 times that observed in the dexamethasone (0.1 mg/mL) positive control group, indicating that its anti-inflammatory activity is comparable to that of dexamethasone (0.1 mg/mL). In the chicken embryo chorionic membrane experiment, it was observed that the safflower fermentation solution did not cause significant damage to the blood vessels of the chorionic allantoic membrane (CAM). This finding demonstrates that the safflower fermentation solution possesses a certain degree of safety.DiscussionSafflower fermentation solution has antioxidant and anti-inflammatory bioactivities, and it has passed cosmetic safety evaluations. It can be used as a new natural cosmetic ingredient added to cosmetic products.

Sustainable bioconversion of rice straw into indole-3-acetic acid by Streptomyces coelicoflavus using response surface methodology

AbstractRice straw is an abundant agricultural waste that poses environmental disposal challenges and can be utilized for biotechnological applications. This study investigates the potential of actinobacteria to enhance rice straw biodegradation and sustainable indole-3-acetic acid (IAA) production, addressing the need for sustainable agricultural practices. Certain actinobacteria strains can effectively degrade rice straw while optimizing IAA production under controlled fermentation conditions. Twenty actinobacteria isolates were screened for lignocellulolytic enzyme activity, and ten were selected for rice straw biodegradation. IAA production was further optimized using response surface methodology based on temperature, pH, and agitation speed (RPM). Isolate S16 achieved a degradation rate of 68.75%, while S18 produced the highest IAA concentration (1040.625 μg/mL) under optimized conditions (25 °C, pH 9, 160 RPM). The purified IAA significantly improved Medicago sativa L. growth. Furthermore, the 16S rRNA gene sequence of isolate S18 was identified it as Streptomyces coelicoflavus strain NSH24 with accession number PP 320383.1. These findings underscore the potential of actinobacteria to efficiently convert agricultural waste into valuable bioproducts and promote sustainable farming practices. By transforming rice straw into high-value products like IAA, this approach contributes to a circular economy, offering an environmentally friendly solution for biomass utilization and agricultural sustainability.

Metabolic engineering of Corynebacterium glutamicum for the production of pyrone and pyridine dicarboxylic acids

Environmental concerns from plastic waste are driving interest in alternative monomers from bio-based sources. Pseudoaromatic dicarboxylic acids are promising alternatives with chemical structures similar to widely used petroleum-based aromatic dicarboxylic acids. However, their use in polyester synthesis has been limited due to production challenges. Here, we report the fermentative production of five pseudoaromatic dicarboxylic acids, including 2-pyrone-4,6-dicarboxylic acid (PDC) and pyridine dicarboxylic acids (PDCAs: 2,3-, 2,4-, 2,5-, and 2,6-PDCA), from glucose using five engineered Corynebacterium glutamicum strains. A platform C. glutamicum chassis strain was constructed by modulating the expression of nine genes involved in the synthesis and degradation pathways of precursor protocatechuate (PCA) and the glucose-uptake system. Comparative transcriptome analysis of the engineered strain against wild-type C. glutamicum identified iolE (NCgl0160) as a target for PDC production. Optimized fed-batch fermentation conditions enabled the final engineered strain to produce 76.17 ± 1.24 g/L of PDC. Using this platform strain, we constructed 2,3-, 2,4-, and 2,5-PDCA-producing strains by modulating the expression of key enzymes. Additionally, we demonstrated a previously uncharacterized pathway for 2,3-PDCA biosynthesis. The engineered strains produced 2.79 ± 0.005 g/L of 2,3-PDCA, 494.26 ± 2.61 mg/L of 2,4-PDCA, and 1.42 ± 0.02 g/L of 2,5-PDCA through fed-batch fermentation. To complete the portfolio, we introduced the 2,6-PDCA biosynthetic pathway to an L-aspartate pathway-enhanced C. glutamicum strain, producing 15.01 ± 0.03 g/L of 2,6-PDCA in fed-batch fermentation. The metabolic engineering strategies developed here will be useful for the production of pseudoaromatic chemicals.

Microorganisms in Fermented Foods and Their Effects on the Human Body

In recent years, microorganisms in fermented foods have shown an important role in regulating human health. Studies have shown that probiotics have significant health benefits in fermented foods, such as improving gut health, stimulating immune function, and promoting metabolism, and mental health. Specifically, lactic acid bacteria in yogurt can increase the number of beneficial bacteria in the intestines and reduce the growth of pathogenic bacteria, and yeasts such as Saccharomyces cerevae have multiple health effects on intestinal microbiota protection and anti-inflammation. In addition, various enzymes and metabolites produced by Aspergillus during fermentation process can improve the nutritional value of food. However, although lots of research has been done, the understanding of the microbial mechanism under different fermentation conditions is still insufficient. In this article, the types and influencing factors of microorganisms in yogurt, wine and seasoner were reviewed. The effects of temperature, pH value, fermentation time and other conditions on microbial activity and product quality were analyzed. The results showed that suitable fermentation conditions could significantly increase microbial activity, improve product flavor and health function, and provide a theoretical basis for optimizing the production process of fermented food. The article provides a theoretical basis for improving the fermented food production process, thus helping to improve product quality and market competitiveness. However, this article has some limitations like the lack of understanding of interactions between different microorganisms. Future research should focus on examining the collaborative relationship between these microbes to improve the health benefits and production efficiency of fermented foods.

A respiratory Streptococcus strain inhibits Acinetobacter baumannii from causing inflammatory damage through ferroptosis

BackgroundMicroecological equilibrium is essential for human health. Previous research has demonstrated that Streptococcus strain A, the main bacterial group in the respiratory tract, can suppress harmful microbes and protect the body. In this study, Streptococcus strain D19T was isolated from the oral and pharyngeal cavities of healthy children. Its antibacterial mechanism against Acinetobacter baumannii was examined, as well as its potential to prevent inflammatory damage to cells. We evaluated the effect of the fermentation conditions of D19T on inhibition of Acinetobacter baumannii growth; Isolation and purification of antibacterial active components of strain D19T and molecular mechanism of inhibition of Acinetobacter baumannii; Molecular mechanism of D19T antibacterial protein reversing cellular inflammatory injury induced by Acinetobacter baumannii.ResultsThe supernatant of fermentation broth of Streptococcus D19T was the active component against Acinetobacter baumannii, but the bacteria had no antibacterial activity. The supernatant of D19T fermentation broth was precipitated by (NH4)2SO4 solution, and the protein was the active antibacterial component. After gel filtration chromatography and anion gel filtration chromatography, the molecular weight of antibacterial protein was 53kD. D19T antibacterial protein can improve cell membrane permeability, limit extracellular soluble protein release, inhibit Acinetobacter baumannii biofilm formation, and prevent Acinetobacter baumannii adhesion. Acinetobacter baumannii induces inflammatory damage to respiratory cells via ferroptosis, and the D19T antibacterial protein can counteract this damage, protecting the respiratory tract.ConclusionStreptococcus strain D19T, as a potential probiotic, inhibits the growth of Acinetobacter baumannii and the inflammatory damage of respiratory cells, playing a protective role in human respiratory health.

Aligning fermentation conditions with non-canonical amino acid addition strategy is essential for Nε-((2-azidoethoxy)carbonyl)-L-lysine uptake and incorporation into the target protein

Protein engineering with non-canonical amino acids (ncAAs) holds great promises for diverse applications, however, there are still limitations in the implementation of this technology at manufacturing scale. The know-how to efficiently produce ncAA-incorporated proteins in a scalable manner is still very limited. In the present study, we incorporated the ncAA N6-[(2-azidoethoxy)carbonyl]-L-lysine (Azk) into an antigen binding fragment (Fab) in Escherichia coli. We used the orthogonal pyrrolysyl-tRNA synthetase/suppressor tRNACUAPyl pair from Methanosarcina mazei to incorporate Azk site-specifically. We characterized Azk uptake and Fab production at bench-scale under different fermentation conditions, varying timing and mode of Azk addition, Azk-to-cell ratio and induction time. Our results indicate that Azk uptake is comparatively efficient in the batch phase. We discovered that the time between Azk uptake and inducing its incorporation into the Fab must be kept short, which suggests that intracellular Azk is consumed and/or degraded. The results obtained in this study are an important step towards the development of efficient production methods for Azk-incorporated proteins in E. coli. The developed process is scalable and provides excellent yields of 2.95 mg functionalized Fab per g CDM, which corresponds to 80% of yield obtained with the wild type Fab. We also identified the cellular uptake of Azk being dependent on the physiological state of the cell as a potential bottleneck in production.

High-Density Fermentation of Lactobacillus plantarum P6: Enhancing Cell Viability via Sodium Alginate Enrichment

Lactobacillus plantarum exhibits a wide range of beneficial physiological functions, including maintaining intestinal microbiota balance, reducing serum cholesterol, and promoting digestive health. According to the specific nutrient requirements of Lactobacillus plantarum P6, we investigated the effects of various carbon sources, nitrogen sources, trace elements, growth-promoting substances, as well as the initial pH and inoculum size on the growth of Lactobacillus plantarum P6 under fermentation conditions. The optimal growth conditions for Lactobacillus plantarum P6 were identified to facilitate high-density fermentation in small-scale fermenter production, achieving a cell concentration of 1.03 × 1011 CFU/mL. This resulted in a 2.5-fold increase in bacterial wet weight, and fermentation time was reduced to 12 h when utilizing a specific medium enriched with 0.2% sodium alginate. It is hypothesized that sodium alginate forms a protective film around the bacterial cells, promoting cell aggregation and enhancing self-coalescence, potentially triggering a bacterial community effect. These results provide a basis for the industrial-scale high-density cultivation of Lactobacillus plantarum, offering potential for enhanced biotechnological applications.

Optimization of Fermentation and Biocontrol Efficacy of Bacillus atrophaeus XHG-1-3m2

Biological control plays an increasingly important role in various aspects of modern agriculture and forestry. Identifying biocontrol strains with commercial potential for effective disease management is currently a focal point in biological control research. In this study, Bacillus atrophaeus XHG-1-3m2, a strain with significant biocontrol potential against Wilsonomyces carpophilus causing shot hole disease in wild apricots, was developed. The study determined the antibacterial activity of the fermentation broth, the optimal fermentation medium composition and conditions, and explored its effectiveness in controlling Wilsonomyces carpophilus. The optimal fermentation medium for strain XHG-1-3m2 comprises 12.5 g/L yeast extract, 12.5 g/L soy peptone, 10.0 g/L sodium chloride, 1 g/L ammonium chloride, 1 g/L potassium dihydrogen phosphate, 1 g/L disodium hydrogen phosphate, and 0.5 g/L magnesium sulfate heptahydrate. With an initial pH of 7.0, a liquid volume of 40%, an inoculum volume of 3%, and shaking incubation at 28 °C for 24 h, the viable cell count reached 14 × 109 CFU/mL. In vitro and in vivo tests on leaves revealed that the fermentation broth and the biocontrol biofertilizer derived from this strain inhibited the leaf lesions caused by Wilsonomyces carpophilus on wild apricots, achieving inhibition rates of 94.62% and 82.46%, respectively.

Evaluation of the Physico-chemical and Biological Properties of Fermented Nipa (Nypa fruticans) Sap Under Closed and Open Vessel Fermentation System for Bioethanol Production

Background The potential of Nipa (Nypa fruticans) sap in the Cagayan Region, Philippines, for bioethanol production remains largely underexplored despite its vast expanse. Methods This study investigates the key physico-chemical properties of Nipa sap from Aparri, Cagayan, with a focus on fermentation dynamics and storage conditions to optimize ethanol production. Two fermentation treatments closed and open vessel conditions, were compared to determine the optimal conditions for maximum ethanol yield. Over a 7-day fermentation period, the acetic acid production, yeast activity, sugar content, and ethanol concentration of closed vessel fermented sap (CVFS) and open vessel fermented sap (OVFS) were monitored. Results CVFS demonstrated a higher ethanol yield (7.56%v/v at peak day), characterized by minimized acetic acid accumulation and stable yeast activity. In contrast, OVFS exhibited significant fluctuations in acetic acid and ethanol levels, indicating a lower fermentation efficiency. Conclusion The presence of methanol and high acetic acid content in open-vessel fermentation systems highlights the importance of fermentation conditions to prevent impurities. This study provides valuable insights into optimizing ethanol production from fermenting Nipa sap by employing closed vessel fermentation, which is crucial for sustainable bioethanol production in the Cagayan region.