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Fermentation
Volume 11
Issue 3
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Fermentation, Volume 11, Issue 3 (March 2025) – 53 articles

Cover Story (view full-size image): Blue corn grain is a cereal with high phenolic compound content. Many of these compounds are bound to the cell matrix, however, limiting their bioavailability. Solid-state fermentation (SSF) using Rhizopus oryzae has aided in the effective release of these compounds in other cereals; however, its application in blue maize has not been studied to the same extent. In this study, we develop a SSF process to improve its phenolic content and antioxidant capacity. We outline the fermentation conditions and metabolite analysis process and characterize the phenolic compounds using HPLC-MS. In addition, we evaluate the changes in antioxidant activity throughout the fermentation process. We propose SSF as a sustainable biotechnological strategy to improve the nutraceutical value of blue maize, highlighting its potential in the food and health industries. View this paper
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17 pages, 672 KiB  
Review
Biotechnological Strategies for Ethanol Reduction in Wine
by Bruno Testa, Francesca Coppola, Mariantonietta Succi and Massimo Iorizzo
Fermentation 2025, 11(3), 159; https://doi.org/10.3390/fermentation11030159 - 20 Mar 2025
Viewed by 681
Abstract
In recent years, wine producers have become increasingly interested in partial or total dealcoholisation of wines due to changing consumer preferences, increased attention to health aspects of alcohol consumption, and the impact of climate change on sugar concentration in grapes. Strategies for decreasing [...] Read more.
In recent years, wine producers have become increasingly interested in partial or total dealcoholisation of wines due to changing consumer preferences, increased attention to health aspects of alcohol consumption, and the impact of climate change on sugar concentration in grapes. Strategies for decreasing alcohol content in wines include pre-fermentation, fermentation, and post-fermentation techniques. These approaches allow for the reduction of fermentable sugars before fermentation, limit the alcohol production during fermentation, and separate alcohol after fermentation. From a biotechnological perspective, yeasts play a critical role in alcoholic fermentation, directly influencing the final alcohol content of the product. The use of non-Saccharomyces yeasts in co-inoculation or sequential inoculation with Saccharomyces yeasts represents a promising biotechnological strategy for the reduction of alcohol in wine. Full article
(This article belongs to the Special Issue Recent Advances in Microbial Fermentation in Foods and Beverages)
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17 pages, 1259 KiB  
Article
Fermentation of Light and Dark Bean Coffee Brews with SCOBY and Lactic Acid Bacteria
by Rusen Metin Yildirim
Fermentation 2025, 11(3), 158; https://doi.org/10.3390/fermentation11030158 - 20 Mar 2025
Viewed by 575
Abstract
This study investigated the effects of fermentation with a SCOBY (symbiotic culture of bacteria and yeast) and lactic acid bacteria (LAB) on the physicochemical and sensory properties of coffee brews prepared from light-roasted (LR) and dark-roasted (DR) coffee beans, with and without the [...] Read more.
This study investigated the effects of fermentation with a SCOBY (symbiotic culture of bacteria and yeast) and lactic acid bacteria (LAB) on the physicochemical and sensory properties of coffee brews prepared from light-roasted (LR) and dark-roasted (DR) coffee beans, with and without the addition of spent coffee grounds (SC). Total phenolic content (TPC), total flavonoid content (TFC), antioxidant activities (DPPH and FRAP), caffeine, and individual phenolic acids were analyzed. Fermentation significantly increased TPC and the concentrations of chlorogenic acids (CGAs), particularly in LR samples, with 5-caffeoylquinic acid (5-CQA) as the most abundant phenolic acid. The addition of spent coffee grounds further enhanced TPC and CGA levels, with total CGA concentrations increasing from 1412.32 to 2458.57 mg/L in LR samples and from 519.77 to 586.37 mg/L in DR samples. Fermentation also led to the isomerization of 5-CQA into 3-CQA and 4-CQA, as well as the release of caffeic acid in LAB-fermented samples. Acetic acid production was exclusive to SCOBY-fermented samples, with higher levels in LR samples (6658 mg/L) compared to DR samples (4331 mg/L). In contrast, lactic acid production was observed only in LAB-fermented samples, reaching 6559 mg/L in LR samples with spent coffee grounds. Antioxidant activity varied depending on the assay, with FRAP values decreasing in fermented samples, while DPPH values remained largely unchanged. Sensory evaluation identified the dark-roasted SCOBY-fermented sample with spent coffee grounds (SK) as the most preferred, characterized by balanced flavor and high overall acceptability. These findings highlight the influence of roasting degree, fermentation type, and substrate composition on the bioactive and sensory properties of fermented coffee, providing insights for the development of novel coffee-based fermented beverages with enhanced functional and sensory profiles. Full article
(This article belongs to the Special Issue Precision Fermentation: Applications in the Food and Beverage Industry)
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15 pages, 4227 KiB  
Article
Biofermentation of Wheat Bran by Monascus anka and Production of High Value-Added Dietary Fiber
by Xuefeng Wu, Siqi He, Hongyi Xu, Hui Zhang, Jing Cai, Min Zhang, Dongdong Mu, Zhenhong Li, Xingjiang Li and Lanhua Liu
Fermentation 2025, 11(3), 157; https://doi.org/10.3390/fermentation11030157 - 20 Mar 2025
Viewed by 449
Abstract
In this study, wheat bran was used to prepare dietary fiber by Monascus anka in liquid fermentation. The structural and functional characteristics of wheat bran dietary fiber were analyzed. Scanning electron microscopy and X-ray diffraction analysis indicated that the insoluble dietary fiber matrix [...] Read more.
In this study, wheat bran was used to prepare dietary fiber by Monascus anka in liquid fermentation. The structural and functional characteristics of wheat bran dietary fiber were analyzed. Scanning electron microscopy and X-ray diffraction analysis indicated that the insoluble dietary fiber matrix was disrupted during the liquid fermentation. Infrared spectroscopy and differential scanning calorimetry analysis demonstrated that intramolecular hydrogen bonds were broken and the oligosaccharides increased. The soluble dietary fiber content increased from 10.7 g/100 g to 16.5 g/100 g, which contributed to improvements in the water-holding capacity, oil-holding capacity, and swelling capacity of wheat bran dietary fiber. UV–Vis spectroscopy analysis demonstrated that the M. anka wheat bran fermentation broth (MWFB) mainly contained yellow pigments (236.6 μ mL−1). HPLC-MS spectrometry further showed MWFB contained three known Monascus pigments: monasine (observed. m/z 359.1853 [M+H]+), ankaflavin (observed. m/z 387.2151 [M+H]+), and monascorubrin (observed. m/z 382.2007 [M+H]+). In conclusion, M. anka can make the most use of wheat bran and improve the structure and function of dietary fiber, thereby expanding its application potential in functional food additives, gut microbiota modulation, and low-calorie baked goods. Full article
(This article belongs to the Special Issue Applied Microorganisms and Industrial/Food Enzymes, 2nd Edition)
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16 pages, 3144 KiB  
Article
Improving Organic Acid Secretion of Aspergillus niger by Overexpression C4-Dicarboxylic Acid Transporters
by Yiyang Tan, Shutong Liu, Sheng Wu, Xiaolu Wang, Depei Wang and Xianli Xue
Fermentation 2025, 11(3), 156; https://doi.org/10.3390/fermentation11030156 - 20 Mar 2025
Viewed by 496
Abstract
C4-dicarboxylic acids are essential organic compounds characterized by a four-carbon structure and two carboxyl groups. Their export from cells is mediated by specialized transporter proteins known as C4-dicarboxylic acid transporters (DCTs). The objective of this study was to investigate the specificity of six [...] Read more.
C4-dicarboxylic acids are essential organic compounds characterized by a four-carbon structure and two carboxyl groups. Their export from cells is mediated by specialized transporter proteins known as C4-dicarboxylic acid transporters (DCTs). The objective of this study was to investigate the specificity of six DCTs (DCT1-5 and C4t318) from Aspergillus niger or Aspergillus oryzae, focusing on their role in different production strategies for C4-dicarboxylic acids. The results indicate that compared to the WT strain, overexpressing dct2 or dct3 in A. niger CGMCC NO. 40550 specifically enhances the production of succinic acid, increasing its yield from 5.69 g/L to 6.28 g/L, and L-malic acid, increasing its yield from 11.02 g/L to 12.11 g/L. Additionally, dct5 appears to be involved in the transport of both succinic acid (6.19 g/L) and L-malic acid (16.33 g/L). The total acid yields of T-D3-7, which lacks the oxaloacetate hydrolase gene, were improved to 27.75 g/L, compared to 25.19 g/L for T-D3-26, due to blocking the branch of oxaloacetate metabolism. Furthermore, the heterologous expression of A. oryzae C4T318 in A. niger increased the production of L-malic acid by approximately 22.5%. Furthermore, the best results were observed when the strains T-D3-7 and T-D5-16 were scaled up in a 30 L bioreactor for 84 h. The succinic acid and L-malic acid yields of T-D3-7 and T-D5-16 reached 14.51 g/L and 70.79 g/L or 41.59 g/L and 81.08 g/L, respectively. Moreover, the purity of L-malic acid produced by T-D3-7 reached 71%. This work further clarifies the specificity of C4-dicarboxylic acid transporters and provides valuable insights for optimizing organic acid production. Full article
(This article belongs to the Special Issue Metabolic Engineering in Microbial Synthesis)
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18 pages, 1246 KiB  
Article
Role of In-House Enzymatic Cocktails from Endophytic Fungi in the Saccharification of Corn Wastes Towards a Sustainable and Integrated Biorefinery Approach
by Patrísia de Oliveira Rodrigues, Anderson Gabriel Corrêa, Lucas Carvalho Basílio de Azevedo, Daniel Pasquini and Milla Alves Baffi
Fermentation 2025, 11(3), 155; https://doi.org/10.3390/fermentation11030155 - 19 Mar 2025
Viewed by 534
Abstract
The valorization of agri-food wastes can provide value-added products, enzymes and biofuels. For the second-generation ethanol (2G) production, pulps rich in cellulose are desirable in order to release fermentable sugars. This study investigated the homemade biosynthesis of cellulases and hemicellulases via solid-state fermentation [...] Read more.
The valorization of agri-food wastes can provide value-added products, enzymes and biofuels. For the second-generation ethanol (2G) production, pulps rich in cellulose are desirable in order to release fermentable sugars. This study investigated the homemade biosynthesis of cellulases and hemicellulases via solid-state fermentation (SSF) using sugarcane bagasse (SB) and wheat bran (WB) for the growth of endophytic fungi (Beauveria bassiana, Trichoderma asperellum, Metarhizium anisopliae and Pochonia chlamydosporia). Cocktails with high enzymatic levels were obtained, with an emphasis for M. anisopliae in the production of β-glucosidase (83.61 U/g after 288 h) and T. asperellum for xylanase (785.50 U/g after 144 h). This novel M. anisopliae β-glucosidase demonstrated acidophile and thermotolerant properties (optimum activity at pH 5.5 and 60 °C and stability in a wide pH range and up to 60 °C), which are suitable for lignocellulose saccharifications. Hence, the M. anisopliae multi-enzyme blend was selected for the hydrolysis of raw and organosolv-pretreated corn straw (CS) and corncob (CC) using 100 CBU/g cellulose. After the ethanol/water (1:1) pretreatment, solid fractions rich in cellulose (55.27 in CC and 50.70% in CS) and with low concentrations of hemicellulose and lignin were found. Pretreated CC and CS hydrolysates reached a maximum TRS release of 12.48 and 13.68 g/L, with increments of 100.80 and 73.82% in comparison to untreated biomass, respectively, emphasizing the fundamental role of a pretreatment in bioconversions. This is the first report on β-glucosidase biosynthesis using M. anisopliae and its use in biomass hydrolysis. These findings demonstrated a closed-loop strategy for internal enzyme biosynthesis integrated to reducing sugar release which would be applied for further usage in biorefineries. Full article
(This article belongs to the Special Issue Fermentative Production of Valuable Chemicals from Lignocellulosic Biomass)
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14 pages, 3828 KiB  
Article
pH-Dependent Metabolic Regulation in Clostridium ljungdahlii During CO Fermentation
by Ze-Rong Liu, Zhi-Qiong Wen, Jing-Wen Wu, Hui-Peng Gao, Quan Zhang, Lan-Peng Li, Li-Cheng Liu, Qiang Li, Fu-Li Li and Zi-Yong Liu
Fermentation 2025, 11(3), 154; https://doi.org/10.3390/fermentation11030154 - 19 Mar 2025
Viewed by 566
Abstract
Clostridium ljungdahlii is a model acetogenic bacterium utilized for ethanol production from syngas, with its growth and ethanol synthesis being profoundly influenced by fermentation pH. However, the mechanistic basis of this pH-dependent regulation remains poorly understood. In this study, we systematically investigated the [...] Read more.
Clostridium ljungdahlii is a model acetogenic bacterium utilized for ethanol production from syngas, with its growth and ethanol synthesis being profoundly influenced by fermentation pH. However, the mechanistic basis of this pH-dependent regulation remains poorly understood. In this study, we systematically investigated the impact of pH on the growth and metabolic profile of C. ljungdahlii under controlled pH conditions using CO as the sole carbon and energy source. At pH 6.0, C. ljungdahlii consumed around 6.0 M carbon monoxide, producing 413 ± 43 mM acetate, 288 ± 35 mM ethanol, and 17 ± 2 mM 2,3-butanediol, with a maximum optical density (OD) of 15.9. In contrast, at pH 5.3, the strain exhibited enhanced metabolic activity, consuming around 9.6 M carbon monoxide and generating 235 ± 24 mM acetate, 756 ± 26 mM ethanol, 38 ± 4 mM 2,3-butanediol, and 28 ± 7 mM lactate, achieving a maximum OD of 30. This represents an approximate twofold increase in both ethanol production and biomass accumulation compared to pH 6.0. Proteomic and parallel reaction monitoring (PRM) analyses demonstrated that the expression levels of key enzymes in central metabolic pathways were marginally higher at pH 6.0 than at pH 5.3, indicating that the observed physiological enhancements were not attributable to differential enzyme expression but likely stemmed from variations in ATP synthesis efficiency. Further optimization experiments revealed that the optimal pH for growth and ethanol production by C. ljungdahlii under CO-sufficient and nutrient-replete conditions is approximately 5.3. These findings provide critical insights into the pH-dependent metabolic regulation of C. ljungdahlii and establish essential parameters for scaling up syngas fermentation for ethanol production. Additionally, this study offers a foundation for further exploration of the unique proton motive force-driven ATP synthesis system in C. ljungdahlii and its broader implications for metabolic network regulation. Full article
(This article belongs to the Special Issue Biofuels Production and Processing Technology, 3rd Edition)
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11 pages, 2878 KiB  
Communication
Microbial Synthesis of Neo-Allo-Ocimene by Celery-Derived Neo-Allo-Ocimene Synthase
by Zheng Liu, Ting Gao, Shaoheng Bao, Penggang Han, Ge Yao, Tianyu Song, Longbao Zhu, Chang Chen and Hui Jiang
Fermentation 2025, 11(3), 153; https://doi.org/10.3390/fermentation11030153 - 18 Mar 2025
Viewed by 411
Abstract
Neo-allo-ocimene is a monoterpene which could be applied in pesticides, fragrances, and sustainable polymers. In this study, we mined a terpene synthase, AgTPS40, from the transcriptome of celery leaf tissues. Through sequence and phylogenetic analysis, AgTPS40 was characterized as a monoterpene synthase. [...] Read more.
Neo-allo-ocimene is a monoterpene which could be applied in pesticides, fragrances, and sustainable polymers. In this study, we mined a terpene synthase, AgTPS40, from the transcriptome of celery leaf tissues. Through sequence and phylogenetic analysis, AgTPS40 was characterized as a monoterpene synthase. The AgTPS40 gene was introduced into a heterologous mevalonate pathway hosted in Escherichia coli to enable terpene production. Gas chromatography–mass spectrometry analysis confirmed that AgTPS40 catalyzes the formation of neo-allo-ocimene, marking the first reported identification of a neo-allo-ocimene synthase. Subsequently, we optimized the fermentation conditions and achieved a yield of 933.35 mg/L in a 1 L shake flask, which represents the highest reported titer of neo-allo-ocimene to date. These results reveal the molecular basis of neo-allo-ocimene synthesis in celery and provide a sustainable way to obtain this compound. Full article
(This article belongs to the Section Microbial Metabolism, Physiology & Genetics)
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20 pages, 3217 KiB  
Article
Enhancement of Biomethane Yield from Spent Mushroom Substrate: Biological Pretreatment with the Chlamydospores of Trichoderma viride
by Wentao Zhu, Xianzhi Lai, Changfa Liu, Xiao Wu, Xiaochen Bai, Yafan Cai, Xiaoling Zhao, Zhe Li, Yongren Hao, Yanhua Huang, Zehui Zheng and Jie Chu
Fermentation 2025, 11(3), 152; https://doi.org/10.3390/fermentation11030152 - 18 Mar 2025
Viewed by 433
Abstract
Fungal chlamydospores are asexual spores formed by fungi under adverse conditions and could be used in biological pretreatment for biogas projects fed by lignocellulosic substrates. In this study, Trichoderma viride (Tv) chlamydospores were used as the pretreatment agent to enhance the methane yield [...] Read more.
Fungal chlamydospores are asexual spores formed by fungi under adverse conditions and could be used in biological pretreatment for biogas projects fed by lignocellulosic substrates. In this study, Trichoderma viride (Tv) chlamydospores were used as the pretreatment agent to enhance the methane yield of spent mushroom substrates (SMSs). Lignocellulosic composition, methanogenesis performance, and anaerobic microbial communities were investigated for different Tv pretreatment durations (0 h, 12 h, 24 h, 48 h, 96 h, and 192 h). The results showed that the optimal Tv pretreatment duration was 24 h, and the cumulative methane yield reached 173.4 mL/gVS, which was 16.8% higher than that of the control. A pretreatment duration longer than 48 h was not conducive to methanogenesis. Sequencing analysis of anaerobic microbial communities showed that the pretreatment duration was directly proportional to the relative abundance of Tv at the beginning of digestion. When the initial Tv abundance was higher than 50%, Trichoderma became the absolute dominant fungus with an abundance higher than 97% in fungal communities in the later stage of digestion. The correlation network among fungi, bacteria, and archaea showed that Tv was directly related to 11 genera, and through these taxa, Tv affected 58% of the taxa in the whole microbial network. Cost accounting showed that Tv pretreatment has a net income of 45.5 CNY/1000 kg SMS, and is a promising technology. This study provides important guidance for the use of fungal chlamydospores in pretreatment and also promotes the understanding of fungi in anaerobic digestion. Full article
(This article belongs to the Special Issue Lignin: Fermentation and Biorefinery Potential)
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21 pages, 9376 KiB  
Article
Optimizing the Production of High-Quality Silage from Jingkenuo 2000 Fresh Waxy Maize: The Synergistic Effects of Microbial Fertilizer and Fermentation Agents
by Min Hou, Yang Li, Fan Yang, Weidong Cui, Xinxin Huang, Deli Dong, Lifeng Dong and Bo Zhang
Fermentation 2025, 11(3), 151; https://doi.org/10.3390/fermentation11030151 - 17 Mar 2025
Viewed by 485
Abstract
Probiotics improve the quality of silage during the planting and fermentation processes. This study was designed to investigate the accumulation of nutritional components in the fresh corn variety Jingkenuo 2000 waxy maize under different fertilization conditions and its effectiveness as silage feed. The [...] Read more.
Probiotics improve the quality of silage during the planting and fermentation processes. This study was designed to investigate the accumulation of nutritional components in the fresh corn variety Jingkenuo 2000 waxy maize under different fertilization conditions and its effectiveness as silage feed. The nutrient accumulation trends of dry matter (DM), starch, neutral detergent fiber (NDF), acid detergent fiber (ADF), crude protein (CP), and ether extract (EE) in the stems, leaves, grains, and whole plant of Jingkenuo 2000 waxy maize were evaluated during different growth cycles. The relative feed value (RFV) was also assessed, with the results showing that the whole plant of Jingkenuo 2000 waxy maize at the wax stage was most suitable for use as silage. The experiment conducted in the present study was carried out in a completely random block design with two additives and three fertilizer application conditions: no-additive + conventional fertilization treatment (CKCK); no-additive + conventional fertilization + drip irrigation of bacterial solution treatment (CKJJ); no-additive + conventional fertilization + microbial organic fertilizer treatment (CKYJ); additive + conventional fertilization treatment (FJCK); additive + conventional fertilization + drip irrigation of bacterial solution treatment (FJJJ); and additive + conventional fertilization + microbial organic fertilizer treatment (YJFJ). Additionally, the nutrient composition, fermentation quality, and bacterial community structure of the silage fermentation treatments were analyzed. The results indicate that there was significant interaction between the additive and fertilization treatments, with them significantly influencing the parameters CP, EE, NDF, ADF, and RFV (p < 0.01). In particular, the treatment combining additives, conventional fertilization, and drip irrigation of bacterial solution exhibited the highest CP, EE, and starch (p < 0.01) among all the tested treatments, while also displaying the lowest NDF and ADF contents (p < 0.01). Furthermore, this treatment reduced the pH value (p < 0.01), decreased bacterial diversity, and fostered the growth of Lactobacillus. Overall, the findings presented herein demonstrate that, through precise nutritional accumulation monitoring and scientific biological pretreatment methods, Jingkenuo 2000 waxy maize has the potential to become a high-quality silage feed. Full article
(This article belongs to the Section Industrial Fermentation)
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23 pages, 827 KiB  
Systematic Review
A Global Review of Geographical Diversity of Kefir Microbiome
by Jeferson Aloísio Ströher, Wemerson de Castro Oliveira, Anderson Santos de Freitas, Marcela Mendes Salazar, Lilian de Fátima Ferreira da Silva, Laís Bresciani, Simone Hickmann Flôres and Patrícia da Silva Malheiros
Fermentation 2025, 11(3), 150; https://doi.org/10.3390/fermentation11030150 - 17 Mar 2025
Cited by 1 | Viewed by 1015
Abstract
Kefir, a traditional probiotic beverage with significant cultural, social, and health relevance, has garnered increasing scientific interest for its functional properties. Here, we synthesized findings from 14 studies investigating the bacterial and fungal diversity in artisanal cow’s milk kefir through metagenomic analysis. Following [...] Read more.
Kefir, a traditional probiotic beverage with significant cultural, social, and health relevance, has garnered increasing scientific interest for its functional properties. Here, we synthesized findings from 14 studies investigating the bacterial and fungal diversity in artisanal cow’s milk kefir through metagenomic analysis. Following the PRISMA guidelines (Preferred Reporting Items for Systematic Reviews and Meta-Analyses), a comprehensive search was conducted in databases including Portal BVS, Scopus, Scielo, and Web of Science. From an initial pool of 522 articles, 14 were selected based on stringent inclusion and exclusion criteria, focusing on English-written studies. Key terms such as “kefir milk”, “artisanal kefir”, “milk”, “metagenomics”, and “cow” were identified through Boolean searches over the last five years. This review addresses the growing need for research on the microbial diversity of artisanal cow’s milk kefir from various global regions. The results indicate a remarkable diversity in microbial communities, primarily dominated by bacteria from the phylum Firmicutes (notably Lactobacillus) and yeasts from the genera Saccharomyces and Kluyveromyces. These microbial compositions are shaped by factors such as milk type, production methods, and grain handling practices, reflecting regional adaptations and influencing kefir’s sensory, probiotic, and functional properties. We argue that a full understanding of these microbial dynamics is critical for standardizing production processes and enhancing quality control measures, ultimately ensuring artisanal kefir’s consistency and health benefits. Full article
(This article belongs to the Special Issue Fermented Dairy Products: From Artisanal Production to Functional Products and Beyond)
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15 pages, 3194 KiB  
Article
Physiological Function of AtrN in Regulating Intracellular NADPH Levels and the Anti-Reductive Stress Response in Corynebacterium glutamicum
by Guotao Xu, Shuping Tian, Zhihan Gong and Jianzhong Xu
Fermentation 2025, 11(3), 149; https://doi.org/10.3390/fermentation11030149 - 17 Mar 2025
Viewed by 471
Abstract
The regulation of intracellular NADPH levels is currently a hotspot for research into bacterial modification and fermentation process optimization, and Corynebacterium glutamicum, an important industrial microorganism, achieves enhanced L-lysine production by regulating intracellular NADPH levels. In previous studies, transcriptome analysis was performed [...] Read more.
The regulation of intracellular NADPH levels is currently a hotspot for research into bacterial modification and fermentation process optimization, and Corynebacterium glutamicum, an important industrial microorganism, achieves enhanced L-lysine production by regulating intracellular NADPH levels. In previous studies, transcriptome analysis was performed on C. glutamicum with different intracellular NADPH levels. The results showed that the expression level of transcription factor AtrN changed significantly. Moreover, experiments showed that transcription factor AtrN can sense high intracellular levels of NADPH and negatively regulate its synthesis. In this study, we integrated the pntAB gene of Escherichia coli into the genome of C. glutamicum XQ-5, successfully constructing a chassis cell with a high intracellular NADPH level. It was named TQ-1. On this basis, we knocked out and complemented the AtrN in strain TQ-1, resulting in strains TQ-2 and TQ-3, respectively. Then, the changes in cell growth, intracellular redox substances and cell membrane among these three strains were investigated. We found that the growth of TQ-2 was inhibited in the early growth stage and the cell survival rate was decreased because of the high increase in the intracellular NADPH level. In addition, the deletion of the AtrN gene also led to a decrease in the fluidity and an increase in the permeability of the cell membrane. Compared with TQ-1, TQ-3 showed slow growth only in the late growth stage, and the fluidity of its cell membrane was also enhanced. This indicates that AtrN guides the cells to make some adaptive changes to maintain cell growth when facing excessive intracellular reductive stress. This will facilitate future research on how potential upstream regulatory genes regulate AtrN and how AtrN regulates downstream genes to cope with cellular reductive stress. It also provides theoretical guidance for the specific modification of high-yield lysine-producing strains. Full article
(This article belongs to the Special Issue New Research on Strains Improvement and Microbial Biosynthesis, 2nd Edition)
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13 pages, 1865 KiB  
Article
Synergistic Effect Enhances Aromatic Profile in Beer Brewing Through Mixed-Culture Fermentation of Pichia kluyveri and Saccharomyces cerevisiae var. diastaticus
by Youyan Rong, Xiaoxue Yu and Kai Hong
Fermentation 2025, 11(3), 148; https://doi.org/10.3390/fermentation11030148 - 17 Mar 2025
Viewed by 562
Abstract
Saccharomyces cerevisiae is one of the important species of traditional fermented foods and beverages. However, incorporating non-Saccharomyces in the fermentation process is a promising strategy to improve the organoleptic profile. In this study, we assessed the potential of a wild Pichia kluyveri [...] Read more.
Saccharomyces cerevisiae is one of the important species of traditional fermented foods and beverages. However, incorporating non-Saccharomyces in the fermentation process is a promising strategy to improve the organoleptic profile. In this study, we assessed the potential of a wild Pichia kluyveri strain (PKL) to augment the aromatic profile in beer brewing while maintaining high fermentation attenuation through inoculation with Saccharomyces cerevisiae var. diastaticus yeast (SY) in both simultaneous (SI-PKL/SY) and sequential (SE-3-PKL/SY) approaches. The fermentation performance was analyzed by residual sugar content, volatile organic compounds, and sensory evaluation. The results indicated that both co-fermentation methods yielded residual sugar levels comparable to those of SY monoculture fermentation. The 2-phenethyl acetate, isoamyl acetate, and linalool in SE-3-PKL/SY increased 12.00, 12.37, and 1.17 folds than the SY monoculture, respectively. Furthermore, the incremental concentrations of these compounds contributed to the highest acceptability and prominent fruity notes in the SE-3-PKL/SY coculture. The current study is the first to report on the co-fermentation with Pichia kluyveri and Saccharomyces cerevisiae var. diastaticus in beer brewing. These findings highlighted the importance of Pichia kluyveri in shaping the ameliorative aroma profile of fermentation production. Full article
(This article belongs to the Special Issue Wine Aromas: 2nd Edition)
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11 pages, 3837 KiB  
Article
Carnosic Acid Production from Sugarcane Syrup by Engineered Yeast in Fed-Batch Fermentation
by Erdem Carsanba, Sara Fernandes, Felipe Beato, Luís Carlos Carvalho, Ana Pintado, Ana Lopes, Mónica Ribeiro, Tânia Leal, Manuela Pintado and Carla Oliveira
Fermentation 2025, 11(3), 147; https://doi.org/10.3390/fermentation11030147 - 15 Mar 2025
Viewed by 662
Abstract
Phenolic diterpene carnosic acid (CA) is widely used in the food, nutritional health, and cosmetic industries due to its antioxidative and antimicrobial properties. This work aimed to overproduce CA in Saccharomyces cerevisiae from sugarcane syrup in fed-batch 2 L bioreactor fermentation. A geranylgeranyl [...] Read more.
Phenolic diterpene carnosic acid (CA) is widely used in the food, nutritional health, and cosmetic industries due to its antioxidative and antimicrobial properties. This work aimed to overproduce CA in Saccharomyces cerevisiae from sugarcane syrup in fed-batch 2 L bioreactor fermentation. A geranylgeranyl diphosphate (GGPP)-producing strain modified with genes encoding the enzymes copalyl diphosphate synthase (Pv.CPS), miltiradiene synthase (Ro.KSL2), hydroxy ferruginol synthase (Ro.HFS), CA synthase (Ro.CYP76AK8), CYP reductase (At.ATR1), and transketolase (TKL1) was used. Lowering the feed rate from 12–26 g/L/h to 7–8 g/L/h, and the use of a dynamic dissolved oxygen (DO) trigger (min. 10%, max. 40%, threshold 70%) instead of a DO trigger of 30%, enhanced CA production by 27%. As a result, the highest CA titer ever reported to date, 191.4 mg/L, was obtained in 4-day fermentation. This study shows the feasibility of engineered yeast to produce CA from the sustainable feedstock sugarcane syrup. Full article
(This article belongs to the Special Issue Current Research in Yeast Fermentation: Physiology, Biotechnology, and Bioprocesses)
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12 pages, 12247 KiB  
Article
Characterization of Low pH and Inhibitor Tolerance Capacity of Candida krusei Strains
by Hironaga Akita, Daisuke Moriguchi and Akinori Matsushika
Fermentation 2025, 11(3), 146; https://doi.org/10.3390/fermentation11030146 - 14 Mar 2025
Viewed by 545
Abstract
Interest in the production of bioethanol from inedible biomass is growing worldwide because of its sustainable supply and lack of competition with food supplies. Candida krusei (also known as Pichia kudriavzevii or Issatchenkia orientalis) is one of the most suitable thermotolerant yeasts [...] Read more.
Interest in the production of bioethanol from inedible biomass is growing worldwide because of its sustainable supply and lack of competition with food supplies. Candida krusei (also known as Pichia kudriavzevii or Issatchenkia orientalis) is one of the most suitable thermotolerant yeasts used in the simultaneous saccharification and fermentation process for bioethanol production. In the production of bioethanol from lignocellulosic biomass as a feedstock, various environmental conditions occur, and the stress tolerance capacity of C. krusei, especially its low pH and tolerance to inhibitors, limits its practical application. In this study, to select a suitable second-generation bioethanol-producing strain, the tolerance capacity of five available C. krusei strains (NBRC0584, NBRC0841, NBRC1162, NBRC1395 and NBRC1664) was characterized. Spot assay and growth experiment results showed that among the five C. krusei strains, C. krusei NBRC1664 showed superior tolerance capacity for low pH and inhibitors. Furthermore, this strain efficiently produced ethanol from glucose under low pH conditions with or without sulfate. A comparative analysis of the draft genome sequences suggested that Opy2, Sln1 and Cdc24 in the HOG pathway are conserved only in C. krusei NBRC1664, which may contribute to its superior tolerance to low pH levels. Moreover, amino acid sequence alignment showed that aldehyde dehydrogenase family proteins, which catalyze the degradation of cyclic aldehydes, are commonly conserved in C. krusei. In addition, the increased transcription levels in C. krusei NBRC1664 could play a role in its higher tolerance to inhibitors. These results suggest that C. krusei NBRC1664 is a more suitable strain for application in industrial processes for second-generation bioethanol production. Full article
(This article belongs to the Special Issue Biofuels and Green Technology)
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14 pages, 2777 KiB  
Article
Optimizing Fermentation of Morus nigra L. Residues with Schizophyllum commune to Enhance Anthocyanin Release and Anti-Inflammatory Activity via Pyroptosis Pathway Modulation
by Lin Ye, Qin Hu, Ying Lin and Chaowan Guo
Fermentation 2025, 11(3), 145; https://doi.org/10.3390/fermentation11030145 - 14 Mar 2025
Viewed by 515
Abstract
Morus nigra L. is rich in anthocyanins and other active ingredients, but its extraction residues pose a burden on the environment. In the present study, Morus nigra L. extraction residue resource utilization was achieved through liquid fermentation of Schizophyllum commune, with the aim [...] Read more.
Morus nigra L. is rich in anthocyanins and other active ingredients, but its extraction residues pose a burden on the environment. In the present study, Morus nigra L. extraction residue resource utilization was achieved through liquid fermentation of Schizophyllum commune, with the aim of enhancing anthocyanin solubilization and evaluating anti-inflammatory activity. Response surface methodology was used to optimize fermentation parameters and quantify anthocyanin fractions by HPLC. The anti-inflammatory effect was evaluated using the lipopolysaccharide-induced inflammation model of human foreskin fibroblast (BJ cell), and the interaction of cyanidin-3-O-glucoside (C3G) with NLRP3, a key target of the pyroptosis pathway, was resolved by molecular docking. Our results indicated that the optimal conditions (substrate 3.4%, inoculum 9%, time 50 h) enabled the total anthocyanin to reach 85.1 μg/mL, of which the C3G content was elevated to 66.7 μg/mL (release efficiency of 83.9%). The fermented filtrate effectively promoted BJ cell proliferation and inhibited the lipopolysaccharide-induced inflammatory response, with the pyroptosis signaling pathway playing a significant role. Molecular docking confirmed that C3G binds strongly to the NLRP3 protein. This technology provides a new strategy for high-value utilization of Morus nigra L. residues and the development of natural anti-inflammatory drugs. Full article
(This article belongs to the Section Industrial Fermentation)
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12 pages, 1224 KiB  
Article
Spectroscopic Analysis of Selenium Nanoparticles Synthesized by Saccharomyces boulardii for the Production of Craft Beer
by Lourdes González-Salitre, Luis Guillermo González-Olivares, Alexis Alejandro Salazar-Navarro, David Cervantes-García, Dagoberto Durán-Hernández, Ricardo Torres-Ramos, Mary Triny Beleño-Cabarcas and Ulin Antobelli Basilio-Cortes
Fermentation 2025, 11(3), 144; https://doi.org/10.3390/fermentation11030144 - 13 Mar 2025
Viewed by 982
Abstract
Selenium is an essential micronutrient which is found in many foods and beverages in low concentrations. Craft beer, one of the most widely consumed fermented beverages globally, presents a strategic opportunity for selenium intake through organic nanoparticles. This study aimed to confirm the [...] Read more.
Selenium is an essential micronutrient which is found in many foods and beverages in low concentrations. Craft beer, one of the most widely consumed fermented beverages globally, presents a strategic opportunity for selenium intake through organic nanoparticles. This study aimed to confirm the presence of selenium nanoparticles in the fermentation process of an ale-style beer using S. boulardii yeast selenized with Na2SeO3 (74 ppm), through spectroscopic analysis and TEM. The yeast accumulated 5.92 mg/g of dry cell mass, and the beer contained 0.642 mg/g of selenium. UV-VIS detected nanoparticles with a peak at 300 nm and FT-IR at a wavelength of 1398.85 cm−1. The particle size ranged between 74 to 175 nm, with a maximum ζ-potential of −4.2 mV, an electrophoretic mobility of −0.3492 μm × cm Vs−1, and a conductivity of 2.656 mS cm−1. TEM analysis revealed that the nanoparticles exhibited circular/ovoid shapes. The fermentation process, combined with the ingredients used to produce ale-type craft beer, proved to be a feasible method for the biosynthesis of selenium nanoparticles using S. boulardii, offering a reliable option for developing and innovating functional craft beers. Full article
(This article belongs to the Special Issue Fermentation: 10th Anniversary)
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20 pages, 6097 KiB  
Article
Transcriptome Analysis of Sclerotium rolfsii: Unraveling Impact of Glycolytic Pathway on Substrate Utilization and Microbial Polysaccharide Production
by Jia Song, Junfeng Li, Chenrui Zhen, Juan Du, Rui Zhao, Bingqian Fan, Jiayi Hou, Bingning Gao, Yu Zheng, Linna Tu and Min Wang
Fermentation 2025, 11(3), 143; https://doi.org/10.3390/fermentation11030143 - 13 Mar 2025
Viewed by 596
Abstract
Scleroglucan is the extracellular polysaccharide (EPS) produced by Sclerotium rolfsii (S. rolfsii). The low EPS titer and limited substrate utilization of S. rolfsii present significant challenges in the fermentation process, restricting industrial applications of scleroglucan. In this study, we performed a [...] Read more.
Scleroglucan is the extracellular polysaccharide (EPS) produced by Sclerotium rolfsii (S. rolfsii). The low EPS titer and limited substrate utilization of S. rolfsii present significant challenges in the fermentation process, restricting industrial applications of scleroglucan. In this study, we performed a transcriptomic analysis on the mycelium of S. rolfsii fermented with different carbon sources. The key genes involved in polysaccharide biosynthesis (6-phosphofructokinase 1 (PFK1), pyruvate decarboxylase (PDC), aldehyde dehydrogenase (NAD (P)+) (ALDH3), and acetyl-CoA synthase (ACS)) were identified and their roles in the process were investigated. The supplementation of specific precursors—fructose-6-phosphate, pyruvate, aldehydes, and acetate—was shown to enhance both the polysaccharide titer and substrate utilization. By adding precursors, the titer of SEPS produced in a 5 L fermentation tank reached 48.69 ± 3.8 g/L. Notably, the addition of these precursors increased the titer of EPS fermented with sucrose (SEPS) by 65.63% and substrate utilization by 119.3%, while the titer of EPS fermented with lactose (LEPS) rose by 80.29% and substrate utilization rose by 47.08%. These findings suggest that precursor supplementation can effectively improve polysaccharide production and substrate efficiency, thereby minimizing resource waste and environmental impact. Full article
(This article belongs to the Special Issue Metabolic Engineering in Microbial Synthesis)
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27 pages, 2570 KiB  
Review
Expanding Layers of Bacteriocin Applications: From Food Preservation to Human Health Interventions
by Furkan Demirgül, Halil İbrahim Kaya, Redife Aslıhan Ucar, Naciye Afranur Mitaf and Ömer Şimşek
Fermentation 2025, 11(3), 142; https://doi.org/10.3390/fermentation11030142 - 13 Mar 2025
Viewed by 1337
Abstract
Bacteriocins, ribosomally synthesized by bacteria, have long been recognized for their role in ensuring food safety and security due to their antibacterial effects against foodborne pathogens and spoilage bacteria. However, recent advancements have unveiled their expanding potential beyond food applications, with increasing evidence [...] Read more.
Bacteriocins, ribosomally synthesized by bacteria, have long been recognized for their role in ensuring food safety and security due to their antibacterial effects against foodborne pathogens and spoilage bacteria. However, recent advancements have unveiled their expanding potential beyond food applications, with increasing evidence of their efficacy against clinically significant pathogenic bacteria, biofilm formation, viral infections, and even cancer. These emerging discoveries have continuously added new layers to the application of bacteriocins, extending their relevance from food preservation to broader human health interventions. To further harness this expanding potential, various innovative strategies have been developed to overcome traditional limitations associated with bacteriocin use. Instead of directly employing bacteriocins or bacteriocin-producing bacterial cultures, novel approaches, such as incorporating them into films and packaging materials or coupling them with nanoparticles, have demonstrated enhanced effectiveness. In this review, we examine the evolving landscape of bacteriocin applications and shed light on the expanding functional spectrum of bacteriocins for both food safety and human health, although some important challenges and limitations remain. By analyzing the recent literature and innovative technological advancements, we highlight how bacteriocins are continuously evolving, opening new frontiers for their use and reinforcing their significance beyond their conventional roles. Full article
(This article belongs to the Special Issue Antimicrobial Metabolites: Production, Analysis and Application)
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14 pages, 1971 KiB  
Article
Metabolic Engineering of Zymomonas mobilis for Xylonic Acid Production from Lignocellulosic Hydrolysate
by Banrui Ruan, Xiongying Yan, Zhaoqing He, Qiaoning He and Shihui Yang
Fermentation 2025, 11(3), 141; https://doi.org/10.3390/fermentation11030141 - 13 Mar 2025
Viewed by 628
Abstract
Bio-based xylonic acid produced from inexpensive lignocellulosic biomass has enormous market potential and enhances the overall economic benefits of biorefinery processes. In this study, the introduction of genes encoding xylose dehydrogenase driven by the promoter Ppdc into Z. mobilis using a plasmid [...] Read more.
Bio-based xylonic acid produced from inexpensive lignocellulosic biomass has enormous market potential and enhances the overall economic benefits of biorefinery processes. In this study, the introduction of genes encoding xylose dehydrogenase driven by the promoter Ppdc into Z. mobilis using a plasmid vector resulted in the accumulation of xylonic acid at a titer of 16.8 ± 1.6 g/L. To achieve stable xylonic acid production, a gene cassette for xylonic acid production was integrated into the genome at the chromosomal locus of ZMO0038 and ZMO1650 using the endogenous type I-F CRISPR-Cas system. The titer of the resulting recombinant strain XA3 reduced to 12.2 ± 0.56 g/L, which could be the copy number difference between the plasmid and chromosomal integration. Oxygen content was then identified to be the key factor for xylonic acid production. To further increase xylonic acid production capability, a recombinant strain, XA9, with five copies of a gene cassette for xylonic acid production was constructed by integrating the gene cassette into the genome at the chromosomal locus of ZMO1094, ZMO1547, and ZMO1577 on the basis of XA3. The titer of xylonic acid increased to 51.9 ± 0.1 g/L with a maximum yield of 1.10 g/g, which is close to the theoretical yield in a pure sugar medium. In addition, the recombinant strain XA9 is genetically stable and can produce 16.2 ± 0.14 g/L of xylonic acid with a yield of 1.03 ± 0.01 g/g in the lignocellulosic hydrolysate. Our study thus constructed a recombinant strain, XA9, of Z. mobilis for xylonic acid production from lignocellulosic hydrolysate, demonstrating the capability of Z. mobilis as a biorefinery chassis for economic lignocellulosic biochemical production. Full article
(This article belongs to the Special Issue Metabolic Engineering in Microbial Synthesis)
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17 pages, 5030 KiB  
Article
Novel Malolactic Fermentation Starter Formulated Using Native Lactic Acid Bacteria Strains from a Re-Emerging Wine-Growing Region of Argentina—A Pilot Scale Vinification
by Gabriel Alejandro Rivas, Naiquen Elizabeth Flores, Natalia Soledad Brizuela, Andrea Cecilia Guillade, Liliana Carmen Semorile and Lucrecia Delfederico
Fermentation 2025, 11(3), 140; https://doi.org/10.3390/fermentation11030140 - 13 Mar 2025
Viewed by 746
Abstract
Argentina ranks worldwide among the top ten wine producers, known for its diverse terroirs and Malbec as its emblematic varietal. Typically, the winemaking process involves alcoholic fermentation, led by yeasts, and malolactic fermentation (MLF), primarily driven by lactic acid bacteria (LAB). Oenococcus oeni [...] Read more.
Argentina ranks worldwide among the top ten wine producers, known for its diverse terroirs and Malbec as its emblematic varietal. Typically, the winemaking process involves alcoholic fermentation, led by yeasts, and malolactic fermentation (MLF), primarily driven by lactic acid bacteria (LAB). Oenococcus oeni and Lactiplantibacillus plantarum are recognised as the best-adapted LAB species for this process. Our previous research focused on a winery located in the southwest of Buenos Aires Province, a scarcely studied re-emerging region of Argentina, which showed a low relative abundance of LAB and incomplete MLF in various vintages. The current study involved the isolation, identification, typing, and use of native strains from the above-mentioned region to formulate a malolactic fermentation starter (MLFS) and to evaluate the strains’ malolactic performance at pilot-scale, implantation capacity and impact on wine aromatic profiles using HS-SPME-GC-FID/MS. Two selected autochthonous strains (Lpb. plantarum UNQLp1001 and a O. oeni UNQOe1101) from the re-emerging region successfully implanted in Malbec wine, achieving faster and more efficient MLF compared to spontaneous MLF. Moreover, the MLFS seems to have influenced the aromatic profile, reducing relative concentrations of alcohols, contributing to the decrease in the bitter and herbaceous notes, and increasing some esters (ethyl acetate, 2-phenethyl acetate, ethyl octanoate), that could enhance floral and fruity, notes. Expanding the availability of candidate strains to formulate native MLFS is a crucial technological tool for the wine industry. Thus, we propose the use of Lpb. plantarum UNQLp1001 and O. oeni UNQOe1101 as potential MLFS in Malbec wines from somewhat similar wine-producing regions. Additionally, the local winery can access a cost-effective MLFS with native LAB strains, enabling a more controlled MLF that preserves regional typicity. Moreover, these strains could enable technology transfer, potentially becoming the first malolactic starters in the region. Full article
(This article belongs to the Special Issue Precision Fermentation: Applications in the Food and Beverage Industry)
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14 pages, 3149 KiB  
Article
Construction of Efficient Multienzyme Cascade Reactions for D-Tagatose Biosynthesis from D-Fructose
by Peiyu Miao, Qiang Wang, Kexin Ren, Tongtong Xu, Zigang Zhang, Runxin Hu, Meijuan Xu, Zhiming Rao and Xian Zhang
Fermentation 2025, 11(3), 139; https://doi.org/10.3390/fermentation11030139 - 12 Mar 2025
Viewed by 674
Abstract
D-tagatose is an ideal sucrose substitute with potential applications in food and healthcare. The combined catalysis of polyphosphate kinase (PPK), fructose kinase (FRK), D-tagatose-6-phosphate 3-differential anisomerase (FbaA) and phytase provides a low-cost and convenient pathway for the biosynthesis of D-tagatose from D-fructose; however, [...] Read more.
D-tagatose is an ideal sucrose substitute with potential applications in food and healthcare. The combined catalysis of polyphosphate kinase (PPK), fructose kinase (FRK), D-tagatose-6-phosphate 3-differential anisomerase (FbaA) and phytase provides a low-cost and convenient pathway for the biosynthesis of D-tagatose from D-fructose; however, there is still a problem of low catalytic efficiency that needs to be solved urgently. Therefore, this study enhanced the biosynthesis of D-tagatose by optimizing the expression levels of PPK, FRK and FbaA in a polycistronic system and knocking out the gene pfka of Escherichia coli. With 30 g/L D-fructose as a substrate, the conversion rate increased to 52%, which was the highest after 24 h. In addition, by constructing a multienzyme self-assembly system with SpyTag and SpyCatcher to improve the whole-cell catalytic ability, the conversion rate was further increased to 75%. Finally, through the fed-batch strategy, the optimal strain Ec-7 produced 68.1 g/L D-tagatose from 100 g/L D-fructose. The multienzyme cascade route reported herein provides an efficient and elegant innovative solution for the generation of D-tagatose. Full article
(This article belongs to the Special Issue 10th Anniversary of Fermentation: Feature Papers in Section “Microbial Metabolism, Physiology & Genetics”)
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19 pages, 7671 KiB  
Article
High-Throughput Sequencing Reveals the Microbial Community Succession Process During the Fermentation of Traditional Daizhou Huangjiu
by Linhua Cui, Jiaying Zhu, Ting Zhang, Qi Li, Yunlong Li and Guoqiang Gao
Fermentation 2025, 11(3), 138; https://doi.org/10.3390/fermentation11030138 - 12 Mar 2025
Viewed by 652
Abstract
This study analyzed the succession patterns of a microbial community structure and key genera during the fermentation process of Daizhou Huangjiu (DZHJ). The results indicated that bacterial diversity decreased while fungal diversity increased in the traditional DZHJ fermentation process. Bacillota and Proteobacteria were [...] Read more.
This study analyzed the succession patterns of a microbial community structure and key genera during the fermentation process of Daizhou Huangjiu (DZHJ). The results indicated that bacterial diversity decreased while fungal diversity increased in the traditional DZHJ fermentation process. Bacillota and Proteobacteria were the dominant bacterial phyla, whereas Ascomycota, Basidiomycota, and Mortierellomycota were the dominant fungal phyla. Weissella, Enterococcus, and Paucibacter were identified as the predominant bacterial genera, with Paucibacter being reported for the first time in Huangjiu research, marking it as a unique signature of DZHJ. Saccharomyces, Aspergillus, and Candida were the predominant fungal genera. Through Beta diversity analysis and LEfSe differential discriminant analysis, Enterococcus, Weissella, and Saccharomyces were confirmed as key differential genera. Additionally, Weissella and Saccharomyces showed significant negative correlations with the majority of bacteria and fungi, respectively. This study elucidates the brewing mechanism of DZHJ, providing a theoretical basis for its quality improvement. Full article
(This article belongs to the Section Probiotic Strains and Fermentation)
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19 pages, 2674 KiB  
Article
Synergistic Optimization of Bacillus subtilis for Efficiently Producing Menaquinone-7 (MK-7) by Atmospheric and Room Temperature Plasma (ARTP) Mutagenesis and Metabolic Engineering
by Meng Li, Jiachang Li, Yufei Li, Xian Zhang and Jianzhong Xu
Fermentation 2025, 11(3), 137; https://doi.org/10.3390/fermentation11030137 - 12 Mar 2025
Viewed by 746
Abstract
Menaquinone-7 (MK-7) plays a crucial role in preventing fractures and certain cardiovascular diseases and is one of the essential vitamins in the human body. In this study, a strain of Bacillus subtilis that produces MK-7 was isolated from commercially available natto fermentation agents, [...] Read more.
Menaquinone-7 (MK-7) plays a crucial role in preventing fractures and certain cardiovascular diseases and is one of the essential vitamins in the human body. In this study, a strain of Bacillus subtilis that produces MK-7 was isolated from commercially available natto fermentation agents, with an MK-7 titer of 75 mg/L. It was named L-5. Firstly, by employing Atmospheric and Room Temperature Plasma (ARTP) mutagenesis technology and protoplast fusion techniques, mutants resistant to 1-hydroxy-2-naphthoic acid (HNA) and diphenylamine (DPA) were obtained, with the titer of MK-7 reaching 196 mg/L. It was named R-8. Based on whole-genome sequencing technology, four mutants involved in the MK-7 synthesis pathway of strain L-5 were identified: 2-succinyl-5-enol-pyruvate-6-hydroxy-3-cyclohexen-1-carboxylic acid, MenD (S249L); (1,4)-dihydroxy-2-naphthalic acid-heptaisoprenyltransferase, MenA (S196L); 1-deoxy-D-xylose-5-phosphate synthetase, Dxs (N60D, Q185H); and hydroxy acid reductive isomerase, Dxr (Q351K). The overexpression of these mutants led to increases in MK-7 production of 19 mg/L, 20 mg/L, 17 mg/L, and 16 mg/L, respectively, compared to the unmutated genes. These mutations have been shown to be effective. To further enhance the production of MK-7, the mutants menD (S249L), menA (S196L), Dxs (N60D, Q185H), and Dxr (Q351K) were co-expressed. The final titer of MK-7 reached 239 mg/L. This study provides theoretical support for the future genetic modification of key enzymes in the MK-7 biosynthetic pathway. Full article
(This article belongs to the Special Issue New Research on Strains Improvement and Microbial Biosynthesis, 2nd Edition)
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2 pages, 327 KiB  
Correction
Correction: Taki et al. Utilization of Okara as a Culture Medium by Membrane Concentration Process for High Oil Production by Oleaginous Yeast, Lipomyces starkeyi. Fermentation 2025, 11, 7
by Hiroya Taki, Kentaro Mine, Mana Miyamoto, Jiro Seto, Shinji Matsuo, Kazuo Kumagai and Hideto Matsuyama
Fermentation 2025, 11(3), 136; https://doi.org/10.3390/fermentation11030136 - 12 Mar 2025
Viewed by 393
Abstract
In the original publication [...] Full article
(This article belongs to the Special Issue Food Wastes: Feedstock for Value-Added Products: 5th Edition)
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17 pages, 1939 KiB  
Article
Artificial vs. Mechanical Daqu: Comparative Analysis of Physicochemical, Flavor, and Microbial Profiles in Chinese Baijiu Starter Cultures
by Huawei Yuan, Zhong Zhang, Liping Ding, Qin Jiang, Qian Li, Jie Huang, Songtao Wang, Li Li, Guohui Nan and Kai Lou
Fermentation 2025, 11(3), 135; https://doi.org/10.3390/fermentation11030135 - 11 Mar 2025
Viewed by 634
Abstract
This study examines the effects of artificial and mechanical production on the physicochemical properties, enzyme activities, flavor components, and microbial diversity of medium-high temperature Daqu, a crucial starter culture for Chinese Baijiu. The research aims to elucidate how these production methods [...] Read more.
This study examines the effects of artificial and mechanical production on the physicochemical properties, enzyme activities, flavor components, and microbial diversity of medium-high temperature Daqu, a crucial starter culture for Chinese Baijiu. The research aims to elucidate how these production methods influence the quality of Daqu and to provide technical insights for optimizing industrial production processes. Results demonstrate that artificial Daqu exhibits 0.24% higher reducing sugar content (p < 0.05), 8.3% greater water retention (p < 0.01), and a 0.10-unit increase in acidity (p < 0.05) compared to mechanically produced Daqu. In contrast, mechanically produced Daqu displays a greater liquefaction capability but reduced fermentation capacity. An analysis of volatile flavor compounds reveals that artificially produced Daqu contains a broader spectrum of aroma compounds, particularly esters and alcohols, which are critical for the flavor profile of Baijiu. Microbial analysis demonstrates that artificially produced Daqu possesses a richer diversity of bacterial and fungal communities, with dominant genera such as Aspergillus, Saccharomyces, and Lactobacillus playing a pivotal role in flavor development. These findings offer valuable insights for optimizing Daqu production methods, balancing traditional craftsmanship with modern industrial demands to ensure consistent quality and flavor in Baijiu production. Full article
(This article belongs to the Special Issue Recent Advances in Microbial Fermentation in Foods and Beverages)
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16 pages, 2921 KiB  
Article
Don’t Judge a Sausage by Its Cover: Effects of Inoculating Three Indigenous Lactic Acid Bacteria on Quality, Moisture Distribution, and Protein Structure in Fermentation
by Yangyi Zheng, Gaiming Zhao, Shichang Zhao, Xuan Li, Wenming Cui, Long Xu, Chaozhi Zhu and Lin Tong
Fermentation 2025, 11(3), 134; https://doi.org/10.3390/fermentation11030134 - 11 Mar 2025
Viewed by 689
Abstract
To produce products with standardized and optimal technical performance, probiotics, particularly Lactic Acid Bacteria (LAB), have long been utilized as fermentation starters in sausages, ensuring both the standardization and enhancement of product quality and safety. Microorganisms isolated from traditional meat products, [...] Read more.
To produce products with standardized and optimal technical performance, probiotics, particularly Lactic Acid Bacteria (LAB), have long been utilized as fermentation starters in sausages, ensuring both the standardization and enhancement of product quality and safety. Microorganisms isolated from traditional meat products, due to their excellent adaptability to the fermentation environment and their ability to preserve desirable flavor, exhibit high potential as candidates for meat fermentation starters. Three indigenous LAB strains—Latilactobacillus sakei, Pediococcus pentosaceus, and Weissella cibaria, isolated from Yunnan ham—were applied in the fermentation of beef sausages to investigate the underlying factors responsible for quality changes. The results indicated that sausages fermented with L. sakei and P. pentosaceus exhibited the lowest pH (4.98) and aw (0.79), while displaying significantly higher hardness, cohesiveness, and chewiness. Additionally, LF-NMR measurements showed that L. sakei and P. pentosaceus promoted the transfer of immobilized water to free water, facilitating the drying and maturation process. Raman spectroscopy analysis revealed a reduction in α-helix content and an increase in disordered β-sheet and β-turn structures in the secondary protein structure. These findings suggest that L. sakei and P. pentosaceus improved quality attributes by modifying the secondary protein structure to enhance water migration and accelerate the ripening process. L. sakei and P. pentosaceus demonstrated desirable technological characteristics, indicating their efficacy for use in fermented sausage production. This study provides valuable insights into improving the production of fermented sausages using specific LAB strains. Full article
(This article belongs to the Section Fermentation for Food and Beverages)
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4 pages, 137 KiB  
Editorial
Technological Innovations and Applications of Microbial Protein Production: From Genetic Engineering to Sustainable Manufacturing
by Yacheng Xu, Yixuan Gao and Dong Liu
Fermentation 2025, 11(3), 133; https://doi.org/10.3390/fermentation11030133 - 10 Mar 2025
Viewed by 811
Abstract
Facing global climate change, resource shortages, and the urgent need for carbon neutrality goals, microbial protein production has demonstrated significant potential in the fields of food, pharmaceuticals, and industrial applications [...] Full article
(This article belongs to the Special Issue Research on Microbial Protein Synthesis)
14 pages, 1025 KiB  
Article
Comparison of Effects of Cold and Warm Water Intake in Winter on Growth Performance, Thermoregulation, Rumen Fermentation Parameters, and Microflora of Wandong Bulls (Bos taurus)
by Jing Li, Zhihao Cui, Ming Wei, Chunqing Yin and Peishi Yan
Fermentation 2025, 11(3), 132; https://doi.org/10.3390/fermentation11030132 - 8 Mar 2025
Viewed by 663
Abstract
Efficient farm practices are crucial for livestock health and performance, and cold stress is a major challenge for cattle in winter. This study aimed to preliminarily investigate the effects of cold and warm water intake in winter on the growth performance, thermal stress [...] Read more.
Efficient farm practices are crucial for livestock health and performance, and cold stress is a major challenge for cattle in winter. This study aimed to preliminarily investigate the effects of cold and warm water intake in winter on the growth performance, thermal stress indicators, serum hormones and metabolites, rumen fermentation parameters, rumen fiber degrading enzyme, and rumen microflora of yellow cattle during winter. Eight Wandong Bulls (Bos taurus) were divided into two groups: group C, which received cold water (6.36 ± 1.99 °C), and group E, which received warm water (32.00 ± 3.12 °C) for 30 d. The results showed that warm water intake significantly increased ADG (p = 0.024) and DMI (p = 0.046) and decreased (p = 0.047) the ratio of feed intake to weight gain. Furthermore, the heat production, respiratory rate, surface temperature, and rectal temperature of cattle did not alter with water temperature, but the heat production value of the bulls increased (29.64 vs. 25.76 MJ/W0.75 h−1) with cold water intake compared to warm water. The concentrations of thyroxine (p = 0.021), serum urea nitrogen (p = 0.025), and glucose (p = 0.011) increased for the bulls drinking cold water compared to those drinking warm water. The concentrations of NH3-N (p = 0.048), total VFA (p = 0.010), acetate (p = 0.009), propionate (p = 0.009), cellulase (p < 0.01), and xylanase (p < 0.05) were lower in group C compared to group E. Total bacterial abundance, as well as specific species including Ruminococcus flavus, Ruminococcus albus, and Prevotella ruminicola, were lower (p < 0.05) in group C compared to group E. In conclusion, drinking warm water during winter enhanced growth performance by influencing energy metabolism, regulating serum hormones and metabolites, and modulating ruminal microflora of bulls compared to cold water intake. Full article
(This article belongs to the Special Issue Ruminal Fermentation)
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12 pages, 1820 KiB  
Article
Metabolic Engineering of Escherichia coli for Xylitol Production
by Jiapeng Li, Lei Zhang, Changzheng Li, Zhaoqing He, Xiongying Yan and Shihui Yang
Fermentation 2025, 11(3), 131; https://doi.org/10.3390/fermentation11030131 - 7 Mar 2025
Viewed by 988
Abstract
Xylitol is a sugar–alcohol compound with broad applications in fields such as the food, dental, and pharmaceutical sectors. Although xylitol biosynthesis has gained attention, the current strategy for industrial xylitol production majorly relies on the chemical hydrogenation of xylose, which is energy-intensive and [...] Read more.
Xylitol is a sugar–alcohol compound with broad applications in fields such as the food, dental, and pharmaceutical sectors. Although xylitol biosynthesis has gained attention, the current strategy for industrial xylitol production majorly relies on the chemical hydrogenation of xylose, which is energy-intensive and environmentally harmful. In this study, the toxicity of xylitol toward Escherichia coli was first examined, and the result demonstrated that Escherichia coli is robust against xylitol at 150 g/L. Genes encoding xylose reductases from different microorganisms were then selected and compared for xylitol production in different E. coli strains. The introduction of xylose reductase of Zymomonas mobilis, driven by the constitutive strong promoter Pgap or Pgap-6M into E. coli, resulted in the accumulation of xylitol at a titer of 64.1 g/L. The increase in NADPH by overexpressing the soluble pyridine nucleotide transhydrogenase encoded by sthA improved the xylitol titer to 83.5 g/L. Seven genes encoding xylose transporters, such as XylE and XylFGH, as well as five mutants of the xylose symporter Glf were then overexpressed and compared for xylitol production. Mutant glfL445I exhibited the highest improvement in xylitol production at a titer of 88.4 ± 0.7 g/L and a yield of 0.95 g/g. Our study thus demonstrated that xylose reductase derived from Z. mobilis is the best one for xylitol production in E. coli, and xylitol production can be further improved by combining diverse metabolic engineering strategies. Our study, thus, provides efficient xylose reductase and a recombinant strain for future industrial xylitol production. Full article
(This article belongs to the Special Issue Microbial Cell Factories for the Production of Functional Compounds)
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Article
Comparative Evaluation of Ensemble Machine Learning Models for Methane Production from Anaerobic Digestion
by Dorijan Radočaj and Mladen Jurišić
Fermentation 2025, 11(3), 130; https://doi.org/10.3390/fermentation11030130 - 7 Mar 2025
Viewed by 761
Abstract
This study provides a comparative evaluation of several ensemble model constructions for the prediction of specific methane yield (SMY) from anaerobic digestion. From the authors’ knowledge based on existing research, present knowledge of their prediction accuracy and utilization in anaerobic digestion modeling relative [...] Read more.
This study provides a comparative evaluation of several ensemble model constructions for the prediction of specific methane yield (SMY) from anaerobic digestion. From the authors’ knowledge based on existing research, present knowledge of their prediction accuracy and utilization in anaerobic digestion modeling relative to individual machine learning methods is incomplete. Three input datasets from compiled anaerobic digestion samples using agricultural and forestry lignocellulosic residues from previous studies were used in this study. A total of six individual machine learning methods and five ensemble constructions were evaluated per dataset, whose prediction accuracy was assessed using a robust 10-fold cross-validation in 100 repetitions. Ensemble models outperformed individual methods in one out of three datasets in terms of prediction accuracy. They also produced notably lower coefficients of variation in root-mean-square error (RMSE) than most accurate individual methods (0.031 to 0.393 for dataset A, 0.026 to 0.272 for dataset B, and 0.021 to 0.217 for dataset AB), being much less prone to randomness in the training and test data split. The optimal ensemble constructions generally benefited from the higher number of individual methods included, as well as from their diversity in terms of prediction principles. Since the reporting of prediction accuracy based on final model fitting and the single split-sample approach is highly prone to randomness, the adoption of a cross-validation in multiple repetitions is proposed as a standard in future studies. Full article
(This article belongs to the Special Issue Current Trends in Bioprocesses for Waste Valorization)
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