High-temperature Daqu Research Articles

Cooperative Interaction Between Pediococcus and Bacillus Communities as a Key Factor in the High-Temperature Thermal Differentiation of Daqu

The high-temperature fermentation process of Daqu, a crucial ingredient in the production of Moutai-flavored Baijiu, is characterized by complex microbial interactions and differentiation at fermentation temperatures. This study is an investigation of the role that microbial cooperation and competition has in driving temperature differences during Daqu fermentation, using the results of real-time temperature monitoring, absolute quantitative amplicon sequencing, and microcalorimetry. The balanced interaction between the genera Pediococcus and Bacillus was observed to be a key factor in achieving high fermentation temperatures. The results indicated that the initial phase of fermentation is characterized by competition and niche differentiation, and that as fermentation proceeds, microbial community of the high-temperature Daqu tends to shift towards cooperation. Correlation network analysis revealed that on the 4th day of fermentation, Pediococcus and Bacillus formed distinct clusters with positive correlations within each cluster and significant negative correlation between them. The interaction and cohesion index analysis suggests that both genera play a crucial role in the high-temperature fermentation process of Daqu. Furthermore, simulated fermentation experiments confirmed that the coexistence of Pediococcus and Bacillus can enhance heat production in high-temperature environments than situations in which either genus dominates. These findings highlight the importance of the microbial community interaction in regulating fermentation temperature and provide valuable insights for optimizing the quality and consistency of Daqu fermentation products.

Generation of a synthetic autochthonous microbiota responsible for the essential flavors and brewing properties of Daqu

Based on the 26 strains of medium–high-temperature Daqu, seed microbial inocula and mature microbial inoculants were prepared and inoculated into crushed wheat to prepare cleaner medium–high-temperature Daqu (XQ). The physicochemical properties, flavor, and microbial profile of XQ were studied using traditional medium–high-temperature Daqu (CQ) or industry standards as controls. The results showed that the liquefaction ability, saccharification activity, esterification activity, and moisture content of XQ were better than those of Daqu industry standards. The results of GC–MS and E-nose analyses showed that the main flavor compounds in the two types of Daqu were esters, alcohols, acids, and aldehydes, which had similar flavor profiles. Microbial sequencing revealed that the most abundant fungi in XQ were Aspergillus subflavus, Wickerhamomyces anomalus, Rhizopus oryzae, and Candida tropicalis, whereas the most abundant bacteria were Lactobacillus curvatus, Weissella cibaria, and Bacillus subtilis. Both types of Daqu contained a high abundance of W. anomalus. In addition, the COG (Clusters of Orthologous Groups of Proteins) analyses displayed that the two types of Daqu had their dominant functions and generally had the pathways and functional characteristics of organism growth and metabolism. This study laid some experimental research bases for the preparation of cleaner Daqu and could provide insights for producing safer microbial fermented foods.

Characterizing the Contribution of Strain Specificity to the Microbiota Structure and Metabolites of Muqu and Fresh High-Temperature Daqu

In this study, the differences in physicochemical properties, microbial community structure, and metabolic characteristics between various fortified Muqu and their corresponding high-temperature Daqu (HTD) were investigated using multiphase detection methods. The results demonstrated that the physicochemical properties, community structure, dominant bacterial composition, and metabolic components varied significantly among the different types of fortified HTD. The differences between HTDs became more pronounced when fortified HTD was used as Muqu. Compared to HTD, Muqu exhibited a more complex volatile profile, while HTD contained higher levels of characteristic non-volatile components. The cultivable bacteria count in Muqu was significantly higher than that in HTD, while the cultivable fungi count was slightly lower than that in HTD. The fungal profiles in HTD were primarily associated with starch hydrolysis and ethanol synthesis, while bacterial activity was more prominent in Muqu. Additionally, pyrazine synthesis was mainly attributed to fungi in Muqu and bacteria in HTD. Source Tracker analysis indicated that 8.11% of the bacteria and 26.76% of the fungi originated from Muqu. This study provides a theoretical foundation for the controlled production of HTD, contributing to improvements in its quality and consistency.

Multi-method joint analysis reveals differences in the quality and microbial composition of high-temperature and medium–high-temperature Daqu

Daqu plays a key role in the fermentation of Baijiu and affects its quality. In this study, Daqu was sourced from a winery in Yingtan City, and the microbial profiles and quality of different Daqu samples were examined using ultra-high-deep metagenomic sequencing and bionic technology, respectively. The starch content of high-temperature Daqu was determined to be 35.65%, which was significantly lower than that of medium–high-temperature Daqu (P < 0.05). In contrast, the acidity, amino nitrogen content, and protein content of high-temperature Daqu were significantly higher than those of medium–high-temperature Daqu (P < 0.05). The fermentation analysis revealed that the alcohol-producing, liquefying, and saccharifying power of high-temperature Daqu were 4.41 U, 2.22 U, and 280.35 U, respectively. These values were all significantly lower than those of medium–high-temperature Daqu (P < 0.05). High-temperature Daqu displayed a higher aroma intensity but weaker bitterness and astringency (P < 0.05). Thus, there were certain differences in sensory quality and fermentation performance between the two types of Daqu. Staphylococcus gallinarum and Kroppenstedtia sp. were mainly found in high-temperature Daqu, while Saccharopolyspora rectivirgula and Saccharopolyspora sp. were found in medium–high-temperature Daqu. Functional analyses revealed that high-temperature Daqu had a higher content of carbohydrate esterases, while medium–high-temperature Daqu had higher auxiliary activity (P < 0.05). The results showed that the temperature-induced differences in the microbial structure of Daqu could affect its sensory quality and fermentation performance. This study provides necessary data support for the screening of wine-making microbes and improvement of Daqu fermentation quality.

Effects of storage period and season on the microecological characteristics of Jiangxiangxing high-temperature Daqu

Metagenomics, non-targeted metabolomics, and metaproteomics were employed to analyze the microecological succession of high-temperature Daqu during storage, elucidate the adaptation mechanism of the microbial community of Daqu to storage environments, and clarify the microecological characteristics of Daqu during different seasons. During storage, the relative abundances of Bacillus, Oceanobacillus, Staphylococcus, and Aspergillus in Daqu had significantly increased, while those of Kroppenstedtia, Saccharopolyspora, Thermoascus, and Thermomyces had significantly decreased. During the first 3 months of storage, compound metabolism of Daqu was primarily dominated by generation of small molecular substances and then shifted to metabolism of amino sugars. During the storage process, homogeneous selection (15.57 %) and homogeneous diffusion (14.86 %) of the microbial communities of Daqu were much larger than during the fermentation process, while the variable selection assembly (29.43 %) was smaller than during the fermentation process. Among the 2509 proteins identified in the four-season Daqu, bacterial protein expression was 1.46-fold greater than that of fungi. Seasonal factors influenced the function of Daqu by alterations to Bacillus subtilis, Oceanobacillus iheyensis, and Aspergillus nidulans and other microbial functions. Carbon and benzoic acid metabolism of Daqu was relatively increased during the spring, while metabolism of alkaloids and tyrosine was upregulated during the summer, amino acid synthesis and starch metabolism were enriched during the autumn, and peptidoglycan synthesis was relatively greater during the winter. Adjusting the moisture content of Daqu during the storage period was shown to reduce microecological differentiation caused by seasonal temperature variations.

Deciphering the effects of different types of high-temperature Daqu on the fermentation process and flavor profiles of sauce-flavor Baijiu

The fermentation of sauce-flavor Baijiu requires high-temperature Daqu as the starter, which is classified into three types: white, yellow, and black. Recent studies highlighted the variability of the microbiota and metabolome profiles of these three types of Daqu, however, what functions they precisely provide in the sauce-flavor Baijiu brewing process remains unclear. In this study, white, yellow, and black Daqu in five different proportions were utilized as starters in field-scale Baijiu fermentation, with the control group lacking Daqu inoculation. Fermented grains inoculated with 100% white Daqu (47.7 °C) had the highest temperature during stacking fermentation, while 100% black Daqu and control groups were lower than 45 °C. Lactic acid (12.6–22.0 mg/g) was the most abundant organic acid, followed by acetic acid (0.7–5.0 mg/g). qPCR analysis revealed that bacterial biomass decreased during fermentation, while fungal biomass increased. A comparison with the control revealed that Daqu contributed high-temperature resistant microorganisms to the stacking fermentation process, primarily Kroppenstedtia, Virgibacillus, Byssochlamys, Thermoascus, and Thermomyces. Meanwhile, stacking fermentation promoted the enrichment of Acetobacter, Kazachstania, and Leiothecium. Mantel test and co-occurrence network analysis revealed that utilizing different fermentation starters could vary the biotic and abiotic factors that drive microbial community succession. We investigated the flavor of Baijiu further, and found that fermentation with white Daqu enhanced the accumulation of ethyl acetate, isoamyl alcohol, and isobutanol, whereas black Daqu increased the accumulation of ethyl lactate, lactic acid, and furfural. This study contributes to the rational usage of different types of Daqu in the manufacturing of Baijiu.

Optimizing the Conditions of Pretreatment and Enzymatic Hydrolysis of Sugarcane Bagasse for Bioethanol Production.

The agricultural waste sugarcane bagasse (SCB) is a kind of plentiful biomass resource. In this study, different pretreatment methods (NaOH, H2SO4, and sodium percarbonate/glycerol) were utilized and compared. Among the three pretreatment methods, NaOH pretreatment was the most optimal method. Response surface methodology (RSM) was utilized to optimize NaOH pretreatment conditions. After optimization by RSM, the solid yield and lignin removal were 54.60 and 82.30% under the treatment of 1% NaOH, a time of 60 min, and a solid-to-liquid ratio of 1:15, respectively. Then, the enzymolysis conditions of cellulase for NaOH-treated SCB were optimized by RSM. Under the optimal enzymatic hydrolysis conditions (an enzyme dose of 18 FPU/g, a time of 64 h, and a solid-to-liquid ratio of 1:30), the actual yield of reducing sugar in the enzyme-treated hydrolysate was 443.52 mg/g SCB with a cellulose conversion rate of 85.33%. A bacterium, namely, Bacillus sp. EtOH, which produced ethanol and Baijiu aroma substances, was isolated from the high-temperature Daqu of Danquan Baijiu in our previous study. At last, when the strain EtOH was cultured for 36 h in a fermentation medium (reducing sugar from cellulase-treated SCB hydrolysate, yeast extract, and peptone), ethanol concentration reached 2.769 g/L (0.353%, v/v). The sugar-to-ethanol and SCB-to-ethanol yields were 13.85 and 11.81% in this study, respectively. In brief, after NaOH pretreatment, 1 g of original SCB produced 0.5460 g of NaOH-treated SCB. Then, after the enzymatic hydrolysis, reducing sugar yield (443.52 mg/g SCB) was obtained. Our study provided a suitable method for bioethanol production from SCB, which achieved efficient resource utilization of agricultural waste SCB.

Species-level understanding of the bacterial community in Daqu based on full-length 16S rRNA gene sequences

Daqu is used as the fermentation starter of Baijiu and contributes diversified functional microbes for saccharifying grains and converting sugars into ethanol and aroma components in Baijiu products. Daqu is mainly classified into three types, namely low (LTD), medium (MTD) and high (HTD) temperature Daqu, according to the highest temperatures reached in their fermentation processes. In this study, we used the PacBio small-molecule real-time (SMRT) sequencing technology to determine the full-length 16 S rRNA gene sequences from the metagenomes of 296 samples of different types of Daqu collected from ten provinces in China, and revealed the bacterial diversity at the species level in the Daqu samples. We totally identified 310 bacteria species, including 78 highly abundant species (with a relative abundance >0.1% each) which accounted for 91.90% of the reads from all the Daqu samples. We also recognized the differentially enriched bacterial species in different types of Daqu, and in the Daqu samples with the same type but from different provinces. Specifically, Lactobacillales, Enterobacterales and Bacillaceae were significantly enriched in the LTD, MTD and HTD groups, respectively. The potential co-existence and exclusion relationships among the bacteria species involved in all the Daqu samples and in the LTD, MTD and HTD samples from a specific region were also identified. These results provide a better understanding of the bacterial diversity in different types of Daqu at the species level.

Comparative analysis of bacterial community structure and physicochemical quality in high-temperature Daqu of different colors in Qingzhou production area

To compare the effects of differences in Daqu making technology and production regions on the bacterial composition and physicochemical properties of high-temperature Daqu (HTD), this study analyzed the bacterial community structure of three colors of HTD in the Qingzhou production area and measured their physicochemical quality. At the same time, a comparative analysis was conducted on the bacterial composition of Qingzhou and Xiangyang regions. The results revealed that the HTD in the Qingzhou area exhibited a diverse bacterial community dominated by Lentibacillus, Scopulibacillus, and Staphylococcus. The black HTD displayed the lowest bacterial richness (P < 0.05) and a relatively unique microbial structure. Significant variations were observed in the physicochemical qualities of the three colors of HTD. Notably, white HTD demonstrated higher moisture and ash content, saccharification and liquor-producing power. Yellow HTD exhibited higher amino nitrogen and protein content, and black HTD displayed higher water activity, acidity, and starch content. The variation in Bacillus, Limosilactobacillus, and Weissella distributions across different colors of HTD primarily contributed to these findings. From the HTD samples in the Qingzhou area, Bacillus (61.90 %) and lactic acid bacteria (17.46 %) being the predominant cultivable communities. Cluster analysis identified significant differences in bacterial communities among HTD samples from various production areas. It can enhance the understanding of HTD quality in the Qingzhou area and offer insights for optimizing HTD and Maotai-flavor Baijiu quality.

Heterogenetic mechanism in high-temperature Daqu fermentation by traditional craft and mechanical craft: From microbial assembly patterns to metabolism phenotypes

The mechanical process has a widely usage in large-scale high-temperature Daqu (HTD) enterprises, however, the quality of the mechanical HTD is gapped with the HTD by traditional process. Currently, the understanding of the mechanism behind this phenomenon is still over-constrained. To this end, the discrepancies in fermentation parameters, enzymatic characteristics, microbial assembly and succession patterns, metabolic phenotypes were compared between traditional HTD and mechanical HTD in this paper. The results showed that mechanical process altered the temperature ramping procedure, resulting in a delayed appearance of the peak temperature. This alteration shifted the assembly pattern of the initial bacterial community from determinism to stochasticity, while having no impact on the stochastic assembly pattern of the fungal community. Concurrently, mechanical pressing impeded the accumulation of arginase, tetramethylpyrazine, trimethylpyrazine, 2-methoxy-4-vinylphenol, and butyric acid, as the target dissimilarities in metabolism between traditional HTD and mechanical HTD. Pearson correlation analysis combined with the functional prediction further demonstrated that Bacillus, Virgibacillus, Oceanobacillus, Kroppenstedtia, Lactobacillus, and Monascus were mainly contributors to metabolic variances. The Redundancy analysis (RDA) of fermented environmental factors on functional ASVs indicated that high temperature, high acid and low moisture were key positive drivers on the microbial metabolism for the characteristic flavor in HTD. Based on these results, heterogeneous mechanisms between traditional HTD and mechanical HTD were explored, and controllable metabolism targets were as possible strategies to improve the quality of mechanical HTD.

Microbial communities, functional, and flavor differences among three different-colored high-temperature Daqu: A comprehensive metagenomic, physicochemical, and electronic sensory analysis

High-temperature Daqu (HTD) is the starter for producing sauce-flavor Baijiu, with different-colored Daqu (white, yellow, and black) reflecting variations in fermentation chamber conditions, chemical reactions, and associated microbiota. Understanding the relationship between Daqu characteristics and flavor/taste is challenging yet vital for improving Baijiu fermentation. This study utilized metagenomic sequencing, physicochemical analysis, and electronic sensory evaluation to compare three different-colored HTD and their roles in fermentation. Fungi and bacteria dominated the HTD-associated microbiota, with fungi increasing as the fermentation temperature rose. The major fungal genera were Aspergillus (40.17%) and Kroppenstedtia (21.16%), with Aspergillus chevalieri (25.65%) and Kroppenstedtia eburnean (21.07%) as prevalent species. Microbial communities, functionality, and physicochemical properties, particularly taste and flavor, were color-specific in HTD. Interestingly, the microbial communities in different-colored HTDs demonstrated robust functional complementarity. White Daqu exhibited non-significantly higher α-diversity compared to the other two Daqu. It played a crucial role in breaking down substrates such as starch, proteins, hyaluronic acid, and glucan, contributing to flavor precursor synthesis. Yellow Daqu, which experienced intermediate temperature and humidity, demonstrated good esterification capacity and a milder taste profile. Black Daqu efficiently broke down raw materials, especially complex polysaccharides, but had inferior flavor and taste. Notably, large within-group variations in physicochemical quality and microbial composition were observed, highlighting limitations in color-based HTD quality assessment. Water content in HTD was associated with Daqu flavor, implicating its crucial role. This study revealed the complementary roles of the three HTD types in sauce-flavor Baijiu fermentation, providing valuable insights for product enhancement.

Pediococcus spp.-mediated competition interaction within Daqu microbiota determines the temperature formation and metabolic profiles.

Fermented microbiota is critical to the formation of microenvironment and metabolic profiles in spontaneous fermentation. Microorganisms generate a diverse array of metabolites concurrent with the release of heat energy. In the case of Daqu fermentation, the peak temperature exceeded 60°C, forming a typical high-temperature fermentation system known as high-temperature Daqu. However, microorganisms that cause the quality variation in Daqu and how they affect the functional microbiota and microenvironment in the fermentation process are not yet clear. This study adopted high-throughput sequencing and monitored the dynamic fluctuations of metabolites and environmental factors to identify the pivotal microorganism responsible for the alterations in interaction patterns of functional keystone taxa and quality decline in the fermentation system of different operational areas during the in situ fermentation process that had been mainly attributed to operational taxonomic unit (OTU)_22 (Pediococcus acidilactici). Additionally, we used isothermal microcalorimetry, plate inhibition experiments, and in vitro simulation fermentation experiments to explore the impact of Pediococcus spp. on heat generation, microorganisms, and metabolite profiles. Results showed the heat peak generated by Pediococcus spp. was significantly lower than that of Bacillus spp., filamentous fungi, and yeast. In addition, the preferential growth of P. acidilactici strain AA3 would obviously affect other strains to colonize through competition, and its metabolites made a significant impact on filamentous fungi. The addition of P. acidilactici strain AA3 in simulated fermentation would cause the loss of pyrazines and acids in metabolites. These evidences showed that the overgrowth of Pediococcus spp. greatly influenced the formation of high temperatures and compounds in solid-state fermentation systems. Our work illustrated the vital impact of interaction variability mediated by Pediococcus spp. for microbial assembly and metabolites, as well as in forming temperature. These results emphasized the functional role of Daqu microbiota in metabolites and heat production and the importance of cooperation in improving the fermentation quality.IMPORTANCEThe stable and high-quality saccharifying and fermenting starter in traditional solid-state fermentation was the prerequisite for liquor brewing. An imbalance of microbial homeostasis in fermentation can adversely impact production quality. Identification of such critical microorganisms and verifying their associations with other fermentation parameters pose a challenge in a traditional fermentation environment. To enhance the quality of spontaneous fermented products, strategies such as bioaugmentation or the control of harmful microorganisms would be employed. This work started with the differences in high-temperature Daqu metabolites to explore a series of functional microorganisms that could potentially contribute to product disparities, and found that the differences in interactions facilitated directly or indirectly by Pediococcus spp. seriously affected the development of microbial communities and metabolites, as well as the formation of the microenvironment. This study not only identified functional microbiota in Daqu that affected fermentation quality, but also demonstrated how microorganisms interact to affect the fermentation system, which would provide guidance for microbial supervision in the actual production process. Besides, the application of isothermal microcalorimetry in this study was helpful for us to understand the heat production capacity of microorganisms and their adaptability to the environment. This study presented a commendable framework for improving and controlling the quality of traditional fermentation and inspired further investigations in similar systems.

Exploring the contribution of temperature-adapted microbiota to enzyme profile of saccharification in Daqu using metagenomics and metaproteomics

Temperature is a critical factor that influences the microbiota in Daqu, which provides essential enzymes for the saccharification process in Baijiu (Chinese liquor) production. However, the relationship between temperature-adapted microbiota and saccharification properties of Daqu remains unclear. In this study, a profile of 23 major saccharifying enzymes and 40 responsible genera was obtained from the metagenomic data of medium- and high-temperature Daqu (MT- and HT-Daqu). The high-temperature pattern (65 °C) primarily affected the saccharification potential of Daqu by promoting Bacillales (fold-change: 0.25–0.75), whereas inhibiting Lactobacillales (fold-change: 3.80–5.60) and Mucorales (fold-change: 5.65–7.09). Eurotiales only exhibited a slight increase (fold-change: 1.01–1.24) to low-temperature pattern (56 °C). Metaproteomic analysis further revealed a notable contribution of Eurotiales in facilitating saccharification. Amylolytic enzymes were primarily derived from Eurotiales species (Byssochlamys spectabilis, Monascus purpureus, Rasamsonia emersonii, and Thermomyces lanuginosus) and one Mucorales species, Lichtheimia romasa. Cellulolytic enzymes were produced by Thermoascus aurantiacus and Paecilomyces sp. Among them, L. romasa was the principal contributor under the low-temperature pattern. Whereas, in HT-Daqu, B. spectabilis, M. purpureus, and T. lanuginosus were the major contributors, as L. romasa is susceptible to higher temperatures. This study offers valuable guidance for targeted optimization of industrial production.

Characterization of key aroma compounds and relationship between aroma compounds and sensory attributes in different quality of high temperature Daqu

Daqu quality is determined by its aroma characteristics. In this study, molecular sensory techniques were used to investigate the sensory attributes of Jiangxiangxing high temperature Daqu, resulting in the development of a sensory wheel. A physicochemical and enzymatic analysis of different grades of Daqu revealed notable differences, especially in acidity. Furthermore, the volatile components of various grades of Daqu were investigated with HS-SPME-GC × GC-TOFMS, resulting in the identification of 524 volatile compounds belonging to 11 distinct classes within Daqu. Univariate analysis demonstrated significant differences in flavor components among Daqu samples of diverse quality grades. By employing GC-O-MS techniques and ROAV≥ 0.1, a comprehensive analysis identified a total of 29 key aroma active compounds that contribute to the overall flavor profile of Daqu. Notably, butyl acrylate and (2E)-2-octenal were found to make substantial contributions to the floral and fruity odor, as well as linalool and benzeneacetaldehyde, which were found to make substantial contributions to sweet odor. These compounds significantly enriched the aroma of Daqu and played a crucial role in enhancing the overall flavor quality of Daqu. In conclusion, these results provide a theoretical base for further investigation of different grades of Daqu, as well as the control and improvement of Daqu quality.

Multidimensional analysis of wheat original crucial endogenous enzymes driving microbial communities metabolism during high-temperature Daqu fermentation

Knowledge of the metabolism of functional enzymes is the key to accelerate the transformation and utilization of raw materials during high temperature Daqu (HTD) manufacturing. However, the metabolic contribution of raw materials-wheat is always neglected. In this research, the relationship between the metabolism of wheat and microorganisms was investigated using physicochemical and sequencing analysis method. Results showed that the process of Daqu generation was divided into three stages based on temperature. In the early stage, a positive correlation was found between Monascus, Rhizopus and glucoamylase metabolism (r > 0.8, p < 0.05). Meanwhile, the glucoamylase metabolism in wheat occupied 63.8 % of the total matrix at the day 4. In the middle to later stages, the wheat metabolism of proteases, α-amylases and lipases in gradually reached their peak. Additionally, Lactobacillus and α-amylases presented a positive correlation (r > 0.7, p < 0.05), and the α-amylases metabolism in wheat occupied 22.18 % of the total matrix during the same time period. More importantly, the changes of enzyme activity metabolic pathway in wheat and microorganism were reflected by respiratory entropy (RQ). Overall, these results guide the choice of substrate during Daqu production.

Combined microbiome and metabolomics analysis of Taorong-type baijiu high-temperature Daqu and medium-temperature Daqu.

Daqu is an essential starter for baijiu brewing in China. However, the microbial enrichment and metabolic characteristics of Daqu formed at different fermentation temperatures are still unclear. High-throughput sequencing technology and the non-targeted metabolomics were used to compare the microbial communities and metabolites of Taorong-type high-temperature Daqu and middle-temperature Daqu. In this study, the relationship between microorganisms and metabolites was established. The study found that the composition and metabolites of the microbial community differed due to the difference in Daqu-making temperature. The bacterial diversity of Taorong-type high-temperature Daqu was higher than that of middle-temperature Daqu, while the fungal community diversity of Taorong-type middle-temperature Daqu was higher than that of high temperature Daqu. A total of 1,034 differential metabolites were screened from the two types of Daqu, and 76 metabolites with significant differences were detected (P < 0.001 and variable importance in projection (VIP) > 1.15). Tetraacetylethylenediamine is the metabolite with the largest differential fold among the 76 differential metabolites, which can be used as a potential marker metabolite of high-temperature Daqu. This study helps elucidate the microbial assembly mechanisms and functional expression under different processing conditions through a further understanding of the composition and metabolic profile differences of different types of Daqu microflora in Taorong-type baijiu.

Wheat-origin Bacillus community drives the formation of characteristic metabolic profile in high-temperature Daqu

The wheat-origin microbiota plays a crucial role in the microbiome assembly of high-temperature Daqu (HTD), particularly during the early incubation stage. In this study, we compared the succession of microbiota and accumulation of metabolites in two groups: one with inactivated wheat-origin microbiotas and the other with non-inactivated microbiotas, under simulated incubation conditions. The results showed that the wheat-origin Bacillus improved the microbial diversity and metabolite profiles by regulating sugar and amino acid metabolisms. Furthermore, four Bacillus strains with excellent fermentation capabilities were specifically isolated and screened from wheat. Bioaugmentation test on HTD further validated that these wheat-origin Bacillus strains enhanced the accumulation of functional enzymes during the early incubation stage. Amylase activity in HTD significantly increased after inoculation with Priestia megaterium, while protease activity saw a significant boost after Bacillus aerophilus inoculation. Additionally, the introduction of wheat-origin Bacillus strains into the HTD microbiome exogenously caused structural changes in the microbial community, heightened the metabolic activity of the microbiota, and led to elevated levels of amino acids and sugars. The collective findings underscore the pivotal role of the wheat-origin Bacillus community in shaping the distinctive metabolic profile of high-temperature Daqu.

Qu-omics elucidates the formation and spatio-temporal differentiation mechanism underlying the microecology of high temperature Daqu

Even if fermented in the same qu-room, Daqu will form various microecologies. A gradual differentiation of microbial population niches was observed within different qu-layers, manifesting as variations in the abundance of dominant microorganisms including Bacillus, Saccharopolyspora, Weissella, Kroppenstedtia, Thermoascus, Thermomyces, Saccharomycopsis, and Rasamsonia. Moreover, distinctions were observed in the functional expression of microorganisms, including Aspergillus, Virgibacillus, Oceanobacillus, and Neurospora. The community in middle layer Daqu exhibited characteristics of high compactness and niche diversity, which facilitated efficient substrate utilization. During the initial phase, the upper Daqu community demonstrated heightened activity. However, in the middle and lower layers, fungi and bacteria respectively exhibited greater functional expression. The key environmental factors regulating the assembly of communities in the upper and middle layers were pH and temperature, respectively, and the lower was moisture and acidity. Notably, deterministic assembly exerted a stronger influence on fungi.

Authenticity identification of high – Temperature Daqu Baijiu through multi-channel visual array sensor of organic dyes combined with smart phone App

High - temperature Daqu Baijiu faces a challenge from illegal adulteration of high-grade Baijiu bottles with low-grade Baijiu, affecting its quality and value. This study developed a rapid identification method for high temperature Daqu Baijiu with the same aroma type using a four-channel visual array sensor and detection of color changes caused by competition coordination with Zn2+ and color-changing organic dyes. The array sensor demonstrated high stability and repeatability in targeting flavor components and achieved 97.78 % or more accuracy combined with DD-SIMCA model in detecting adulteration across the Baijiu with same aroma type. The results of GC–MS and Quantum Chemical Calculation showed that esters, acids, and pyrazines played a crucial role. The smart phone App could quickly identify the authenticity of Baijiu with accuracy achieved 93 %. This research provides a foundation for rapid and reliable assessment of Baijiu quality and authenticity, enabling the industry to combat fraudulent practices effectively.

Solibacillus daqui sp. nov., isolated from high-temperature Daqu.

A Gram-stain-positive, rod-shaped, endospore-forming, aerobic bacterial strain, designated ZS111008T, was isolated from high-temperature Daqu, a starter for production of Chinese Jiang-flavour Baijiu, and was characterized by polyphasic taxonomy. This novel isolate grew in the presence of 0-5 % (w/v) NaCl, at pH 6.0-9.0 and 25-45 °C; optimum growth was observed with 1 % (w/v) NaCl, at pH 8.0 and 30 °C. A comparative analysis of the 16S rRNA gene sequence (1461 bp) of strain ZS111008T showed highest similarity to Solibacillus silvestris DSM12223T (96.7%), followed by Solibacillus cecembensis PN5T (96.6%) and Solibacillus isronensis AMCK01000046 (96.5%). The DNA G+C content of strain ZS111008T was 37.21 mol%. The respiratory quinone was identified as menaquinone-7 and the major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine and one unknown phospholipid. Lys was detected as the diagnostic diamino acid in the cell wall. Based on morphological characteristics, chemotaxonomic characteristics and physiological properties, strain ZS111008T represents a novel species of the genus Solibacillus, for which the name Solibacillus daqui sp. nov. is proposed. The type strain for this proposed species is ZS111008T (=CGMCC 1.19455T=JCM 35214T).