A mutant GH3 family ß-glucosidase from Oenococcus oeni exhibits superior adaptation to wine stresses and potential for improving wine aroma and phenolic profiles.

In this study, we conducted a comprehensive investigation into a GH3 family ß-glucosidase (BGL) from the wild-type strain of Oenococcus oeni and its mutated counterpart from the acid-tolerant mutant strain. Our analysis revealed the mutant BGL's remarkable capacity to adapt to wine-related stress conditions, including heightened tolerance to low pH, elevated ethanol concentrations, and metal ions. Additionally, the mutant BGL exhibited superior hydrolytic activity towards various substrates. Through de novo modeling, we identified specific amino acid mutations responsible for its resilience to low pH and high ethanol environments. In simulated wine conditions, the mutant BGL outperformed both wild-type and commercial BGLs, efficiently releasing terpene and phenolic aglycones from glycosides in wine grapes. These findings not only expand our understanding of O. oeni BGLs but also highlight their potential in enhancing wine production. The mutant BGL's enhanced adaptation to wine stress conditions opens promising avenue for improving wine quality and flavor.

Integrative multiomics analysis of the acid stress response of Oenococcus oeni mutants at different growth stages.

BACKGROUND: Acid stress is one of the most important environmental stresses that adversely affect the growth of lactic acid bacteria (LAB), such as Oenococcus oeni which was isolated from grape-berries and mainly used in wine fermentation. The aim of this paper is to comprehensively characterize the mechanisms of acid stress regulation in O. oeni and to provide a viable theoretical basis for breed and improvement of existing LAB. METHOD: First, six O. oeni mutants with acid-sensitive (strains b2, a1, c2) and acid-tolerant (strains b1, a3, c1) phenotypes were screened from three wild-type O. oeni, and then their genome (sequencing), transcriptome and metabolome (LC-MS/MS) were examined. RESULTS: A total of 459 genes were identified with one or more intragenic single nucleotide polymorphisms (SNPs) in these mutants, and were extensively involved in metabolism and cellular functions with a high mutation rates in purine (46%) and pyrimidine (48%) metabolic pathways. There were 210 mutated genes that cause significant changes in expression levels. In addition, 446 differentially accumulated metabolites were detected, and they were consistently detected at relatively high levels in the acid-tolerant O. oeni mutant. The levels of intracellular differentially expressed genes and differential metabolites changed with increasing culture time. CONCLUSION: The integrative pathways analysis showed that the intracellular response associated with acid regulation differed significantly between acid-sensitive and acid-tolerant O. oeni mutants, and also changed at different growth stages.

Regulated synthesis and metabolism of Monascus pigments in a unique environment.

Microbial fermentation in extreme environments is the focus of research, which provides new insights for the production and application of Monascus pigments. In this paper, the regulation of Monascus pigments metabolism by optimizing the conditions, such as osmotic pressure, high sugar stress, light, extreme temperature, high-water content, low-frequency magnetic field and ultrasonics, is reviewed and summarized in four parts: the mycelium growth of Monascus spp., metabolic pathways, gene expression and composition characteristics of Monascus pigments. The relationship between mycelial morphology, gene expression and pigments production during fermentation under unique environments is discussed. Based on the changes in metabolic pathways and composition characteristics, the regulatory mechanism of Monascus pigments under unique conditions is proposed. Moreover, the fermentation strategy and application prospects of Monascus pigments in unique environments are also discussed. This work will provide a theoretical basis and practical guidance for the optimized production of Monascus pigments.

Your losses are mine: The influence of empathic concern on evaluative processing of others' outcomes.

Neural responses to others' decision-making outcomes can be modulated by many social factors. Using the event-related potential (ERP) technique, we explored the neural mechanisms of empathic concern modulating evaluative processing of others' outcomes. Participants were asked to perform a gambling task for three beneficiaries: themselves and two strangers. One stranger was an economically underprivileged student requiring help (high-empathy condition); the other stranger was a student with no upsetting information to induce empathic concern (low-empathy condition). ERP results showed that the valence effect of the feedback-related negativity (FRN) was larger when participants exhibited high empathic concern than when they did not. The FRN responses to strangers' outcomes in the high-empathy condition were as strong as those to their own outcomes. The P300 showed no differences between the low- and high-empathy conditions. These findings indicate that empathic concern could modulate the early stage of outcome processing, implying empathic emotional/altruistic motivational impacts of others' outcomes.

Investigation of the mycelial morphology of Monascus and the expression of pigment biosynthetic genes in high-salt-stress fermentation.

Extreme environments, for example high-salt-stress condition, that can induce secondary metabolite biosynthesis in fungi are a promising and effective strategy for producing natural Monascus pigments used as food colourants and nutraceutical supplements. In this study, the relationship between the mycelial morphology and expression of pigment biosynthetic genes in high-salt-stress fermentation (HSF) with Monascus ruber CGMCC 10910 was investigated. The Monascus fungus grew well under HSF conditions with 35 g/l NaCl, and the intracellular yellow pigment yield in HSF was 40% higher than that in conventional batch fermentation (CBF). Moreover, the mycelial morphology was maintained in a better state, with a hyphal diameter of 5-6 µm in HSF, indicating good biocatalytic activity for pigment synthesis. The rate of the relative content of intracellular orange pigments to yellow pigments (O/Y) significantly (p < 0.05) changed, and the extracellular yellow pigments were transformed into each other, indicating that the pigment biosynthesis pathway was changed to promote yellow pigment accumulation in HSF. The pigment biosynthesis genes MpPKS5, MpFasB2, mppE, mppD and mppB were significantly (p < 0.05) up-regulated by approximately 58.4-106.1%, whereas the regulatory genes mppR1 and mppR2 were significantly (p < 0.05) down-regulated by approximately 23.2% and 59.0% in HSF. Notably, the mppE gene was highly correlated with (r > 0.95, p < 0.05) hyphal diameter. These findings indicated that the cultivation of the Monascus fungus under high-salt-stress conditions was beneficial for pigment biosynthesis by controlling the mycelial morphology to regulate gene expression. This study first described the relationship between the mycelial morphology and expression of pigment biosynthetic genes in Monascus during fermentation. KEY POINTS: • High-salt-stress fermentation (HSF) was first performed to improve Monascus pigment yield. • Pigment biosynthesis was enhanced by maintaining the mycelial morphology in an improved state in HSF. • Gene expression was up-/downregulated to promote yellow pigment accumulation in HSF. • The mycelial morphology was highly related to the expression of pigment biosynthetic genes in HSF.

Dynamic Changes in Metabolite Accumulation and the Transcriptome during Leaf Growth and Development in Eucommia ulmoides.

Eucommia ulmoides Oliver is widely distributed in China. This species has been used mainly in medicine due to the high concentration of chlorogenic acid (CGA), flavonoids, lignans, and other compounds in the leaves and barks. However, the categories of metabolites, dynamic changes in metabolite accumulation and overall molecular mechanisms involved in metabolite biosynthesis during E. ulmoides leaf growth and development remain unknown. Here, a total of 515 analytes, including 127 flavonoids, 46 organic acids, 44 amino acid derivatives, 9 phenolamides, and 16 vitamins, were identified from four E. ulmoides samples using ultraperformance liquid chromatography-mass spectrometry (UPLC-MS) (for widely targeted metabolites). The accumulation of most flavonoids peaked in growing leaves, followed by old leaves. UPLC-MS analysis indicated that CGA accumulation increased steadily to a high concentration during leaf growth and development, and rutin showed a high accumulation level in leaf buds and growing leaves. Based on single-molecule long-read sequencing technology, 69,020 transcripts and 2880 novel loci were identified in E. ulmoides. Expression analysis indicated that isoforms in the flavonoid biosynthetic pathway and flavonoid metabolic pathway were highly expressed in growing leaves and old leaves. Co-expression network analysis suggested a potential direct link between the flavonoid and phenylpropanoid biosynthetic pathways via the regulation of transcription factors, including MYB (v-myb avian myeloblastosis viral oncogene homolog) and bHLH (basic/helix-loop-helix). Our study predicts dynamic metabolic models during leaf growth and development and will support further molecular biological studies of metabolite biosynthesis in E. ulmoides. In addition, our results significantly improve the annotation of the E. ulmoides genome.

Carbon storage regulator CsrA plays important roles in multiple virulence-associated processes of Clostridium difficile.

The carbon storage regulator CsrA is a global regulator that controls multiple virulence-associated processes including host cell invasion, virulence secretion, quorum sensing, biofilm formation, and motility in many pathogenic bacteria. However, the roles of CsrA in Clostridium difficile still remain unclear. In this study, a C. difficile strain overexpressing csrA was constructed to investigate its effects on multiple virulence associated processes. Overexpression of csrA resulted in flagella defect and poor motility in C. difficile 630Δerm, suggesting that CsrA involves in the regulation of flagellum synthesis. The levels of toxin production were increased in the C. difficile 630Δerm overexpressing of csrA. Moreover, csrA overexpression enhanced the adherence ability to Caco-2 cells and solvent production of C. difficile 630Δerm. Altogether, CsrA of C. difficile participates in multiple virulence processes including toxin production, motility, and adherence, and in the regulation of carbon metabolism. These results enhance our understanding of the regulatory functions of CsrA and reveal that CsrA is an important regulator in C. difficile contributing to virulence regulation.

High-mobility group box 1 protein contributes to the immunogenicity of rTcdB-treated CT26 cells.

Clostridium difficile TcdB is a key virulence factor that causes C. difficile-associated diseases. Our previous studies have shown that recombinant full-length TcdB (rTcdB) induces cell death in CT26 cells, and rTcdB-treated CT26 cells with high immunogenicity could stimulate dendritic cell (DC) activation and T cell activation in vitro. The rTcdB-treated CT26 cells also induce antitumor immunity in mice and protect mice from CT26 cells. High-mobility group box 1 protein (HMGB1) is a non-histone nuclear protein, which has various biological functions within the nucleus and also acts as an extracellular signal molecule involving in inflammatory diseases, cancers or autoimmune diseases. In this study, HMGB1 was found to be released from the rTcdB-treated CT26 cells. HMGB1 knockdown by using specific siRNA weakened the capacity of the BMDCs loaded with the rTcdB-treated CT26 cells to prime T cells in vitro and in vivo. The released HMGB1 from CT26 cells could interact with the receptor TLR4, which is closely related to DC activation and immune responses. The knockdown of HMGB1 also affected the phagocytosis of the rTcdB-treated CT26 cells by DCs in vitro. Furthermore, HMGB1 weakened the antitumor immunity of the rTcdB-treated CT26 cells, which protects mice from rechallenge of the live CT26 cells. Taken together, these results suggest that HMGB1 plays an important role on the immunogenicity of the rTcdB-treated dying CT26 cells.

Pigment fingerprint profile during extractive fermentation with Monascus anka GIM 3.592.

BACKGROUND: Traditional submerged fermentation mainly accumulates intracellular orange pigments with absorption maxima at 470 nm, whereas extractive fermentation of Monascus spp. with Triton X-100 can promote the export of intracellular pigments to extracellular broth, mainly obtaining extracellular yellow pigments with absorption maxima at approximately 410 nm. In this study, a strain of Monascus (M. anka GIM 3.592) that produces high yields of pigments was employed to investigate the differences in pigment fingerprint profiles between submerged and extractive fermentations. RESULTS: Using extractive fermentation with this high-yield strain, the extracellular pigments exhibited an absorption maximum at 430 nm, not 410 nm, as previously observed. By comparing the pigment fingerprint profiles between submerged and extractive fermentations, extractive fermentation was found to not only export intracellular pigments to the extracellular broth, but also to form four other yellow pigments (Y1-Y4) that accounted for a large proportion of the extracellular pigments and that were not produced in submerged fermentation. The yields of Y1-Y4 were closely related to the concentration and feeding time point of Triton X-100. Y1-Y4 presented identical UV-Vis spectra with absorption maxima at 430 nm and fluorescence spectra with absorption maxima (emission) at 565 nm. HPLC-MS and the spectral analysis showed that the four pigments (Y1-Y4) had not been previously reported. The results indicated that these pigments may rely on the bioconversion of orange pigments (rubropunctatin and monascorubrin). CONCLUSIONS: Using extractive fermentation with M. anka led to a high yield of extracellular yellow pigments (AU410 nm = 114), and the pigment fingerprint profile significantly differed compared to the results of traditional submerged fermentation. These results provide information and a detailed view of the composition and variation of pigments in extractive fermentation and could also contribute to characterizing pigment metabolism during extractive fermentation.

Fingerprint profiles of flavonoid compounds from different Psidium guajava leaves and their antioxidant activities.

Flavonoids are the main active components in Psidium guajava leaves and have many multi-physiological functions. In this study, the flavonoid compositions were identified in the Psidium guajava leaves samples using a high-performance liquid chromatography with time-of-flight electrospray ionization mass spectrometry method. A high-performance liquid chromatography fingerprint method, combined with chemometrics, was used to perform a quality assessment of the Psidium guajava leaves samples. The eight identified flavonoid compounds including rutin, isoquercitrin, quercetin-3-O-ß-d-xylopyranoside, quercetin-3-O-α-l-arabinopyranoside, avicularin, quercitrin, quercetin, and kaempferol were used as the chemical markers. The antioxidant activity of 15 batches of samples was examined using three different methods, and the results revealed the Psidium guajava leaves samples that had higher contents of the flavonoid compounds, glycoside and aglycone, possessed the highest antioxidant capacities. Consequently, a combination of chromatographic fingerprints and chemometric analyses was used for a quality assessment of Psidium guajava leaf tea and its derived products, which can lay the foundation for the development of plant tea resources or other herbs.

Chemical Compositions, Antioxidant and Antimicrobial Activities of Essential Oils of Psidium guajava L. Leaves from Different Geographic Regions in China.

Hydrodistilled essential oils (EO) of Psidium guajava L. leaves from different regions in China were analyzed by GC and GC/MS. The samples from Guangdong Province displayed high EO yields (0.61 - 0.75%, v/w). A total of 50 components, representing over 98.00% of the EOs, were identified and semi-quantitatived. The major constituents of EOs included ß-caryophyllene (17.17 - 31.38%), γ-gurjunene (9.17 - 15.22%), τ-cadinol (1.35 - 10.02%) and calamenene (2.13 - 7.80%). The terpenoids in all sample oils were dominated by sesquiterpenes hydrocarbons (70.18 - 84.35%), followed by oxygenated sesquiterpenes (9.89 - 22.19%). The similarities and differences among EOs from different samples were evaluated by hierarchical cluster analysis and principal component analysis methods. The IC50 values of EOs from different regions were between 18.52 - 33.72 mg/ml (DPPH) and 13.12 - 25.15 mg/ml (ABTS+ ). The FRAP value of EO from Guangdong Province was 7.34 - 9.13 mmol Vc/g DM, while the FRAP value of EO from Taiwan Province was 2.29 - 2.36 mmol Vc/g DM. The antimicrobial tests revealed that EO had a higher antimicrobial activity against all Gram-positive bacteria and two fungi. Moreover, EO from P. guajava leaves of Guangdong Province showed the highest antimicrobial activity. These properties can be considered in the design of industrial products and for further application in the food, pharmaceutical and cosmetic industries.

The pigment characteristics and productivity shifting in high cell density culture of Monascus anka mycelia.

BACKGROUND: Monascus mycelia and pigments are promising sources of food and medicine with their potential pharmaceutical values and health-improving functions. Using high cell density fermentation of Monascus spp. to achieve higher mycelium and yellow pigment production is worthy to be researched. In this study, the characteristics and productivity shifting of pigments in high cell density culture of Monascus anka GIM 3.592 were investigated. RESULTS: The high yield of Monascus mycelia up to 39.77 g/L dry cell weight (DCW), which was achieved by fed-batch fermentation with the feeding medium containing C, N, P and trace elements, was four times higher than that of conventional batch culture. But the total pigment production decreased by 14.6 %, which suggested non-coupled growth. Potential novel yellow pigments accumulated constantly at the late stage of the fed-batch culture, which resulted in a shift in pigment characteristics so that yellow pigments became the dominant pigments. Citrinin production was extremely low and independent of feeding ingredients. CONCLUSIONS: This study provided a suitable fermentation strategy to produce functional Monascus mycelia with a high proportion of yellow pigments in high cell density culture. For the first time, it reported the pigment productivity and characteristics shifting in high cell density culture of Monascus.

Design and stability analysis of the gear-type mobile mechanism with a single actuator.

Gear mechanism transmits the motion and power of parallel axes, intersecting axes and staggered axes, which has been widely employed in production and life. Gear-type mobile mechanism is a type of robot that can achieve motion through gear transmission. Due to its unique motion mode, it can handle various tasks in certain unconventional environments, such as particularly steep surfaces. Cylindrical gear, bevel gear and non-cylindrical gear are taken as the main parts of the mechanism to form a novel research series, respectively. The models of gear-type mobile mechanism are established in this paper, and the degrees of freedom of the mechanism are briefly calculated based on the screw theory. Simultaneously, the influence of centroid trajectory on motion stability is discussed to solve the possible problem that opposite rotation occur during the movement. Furthermore, the trajectory model of zero moment point is calculated considering the motion of the gear-type mobile mechanism. After that, the simulation and experimental analysis of its motion capability show that the gear-type mobile mechanism has excellent characteristics in stability, flexibility and continuity.

Foodomics uncovers functional and volatile metabolite dynamics in red raspberry chewable tablet optimized processing.

Raspberries are known to contain valuable metabolites and possess a robust antioxidant capacity. However, the impact of different tablet processing stages on the nutritional content and flavor profile of raspberries remains unclear. The dynamic profile of functional and volatile metabolites was investigated through foodomics combined with UPLC-MS/MS-based widely targeted metabolomics and HS-SPME-GC-MS, and antioxidant capacities were assessed during tablet processing. 1336 functional metabolites and 645 volatile metabolites were identified. Results indicated tablets retained 34% âˆ¼ 61% of the total volatile contents. In addition, the conversion intensity of functional metabolites was consistent with the order of "Tableting > Freeze-drying > Crushing". Compared to raspberry, tablets showed higher antioxidant activity, which was positively correlated with vitamin contents. This study elucidated that tablet formation demonstrated advantages in antioxidation and aroma retention, which may provide insights for enhancing quality during the tableting process.

Impact of indigenous Oenococcus oeni and Lactiplantibacillus plantarum species co-culture on Cabernet Sauvignon wine malolactic fermentation: Kinetic parameters, color and aroma.

Malolactic fermentation (MLF) is a crucial process to enhance wine quality, and the utilization of indigenous microorganisms has the potential to enhance wine characteristics distinct to a region. Here, the MLF performance of five indigenous Oenococcus oeni strains and six synthetic microbial communities (SynComs), were comparatively evaluated in Cabernet Sauvignon wine. In terms of malate metabolism rate and wine aroma diversity, the strain of O. oeni Oe114-46 demonstrated comparable MLF performance to the commercial strain of O. oeni Oe450 PreAc. Furthermore, the corresponding SynComs (Oe144-46/LpXJ25) exhibited improved fermentation properties, leading to increased viable cell counts of both species, more rapid and thorough MLF, and increased concentrations of important aroma compounds, such as linalool, 4-terpinenol, α-terpineol, diethyl succinate, and ethyl lactate. These findings highlight the remarkable MLF performance of indigenous O. oeni and O. oeni-L. plantarum microbial communities, emphasizing their immense potential in improving MLF efficiency and wine quality.

Improving Soluble Phenolic Profile and Antioxidant Activity of Grape Pomace Seeds through Fungal Solid-State Fermentation.

Grape pomace seeds contain abundant phenolic compounds, which are also present in both soluble and insoluble forms, similar to many other plant matrices. To further increase the extractable soluble phenolics and their antioxidant activities, grape pomace seeds were fermented with different fungi. Results showed that solid-state fermentation (SSF) with Aspergillus niger, Monascus anka, and Eurotium cristatum at 28 °C and 65% humidity had a significantly positive impact on the release of soluble phenolics in grape pomace seeds. Specifically, SSF with M. anka increased the soluble phenolic contents by 6.42 times (calculated as total phenolic content) and 6.68 times (calculated as total flavonoid content), leading to an overall improvement of antioxidant activities, including DPPH (increased by 2.14 times) and ABTS (increased by 3.64 times) radical scavenging activity. Furthermore, substantial changes were observed in the composition and content of individual phenolic compounds in the soluble fraction, with significantly heightened levels of specific phenolics such as chlorogenic acid, syringic acid, ferulic acid, epicatechin gallate, and resveratrol. Notably, during M. anka SSF, positive correlations were identified between the soluble phenolic content and hydrolase activities. In particular, there is a strong positive correlation between glycosidase and soluble phenols (r = 0.900). The findings present an effective strategy for improving the soluble phenolic profiles and bioactivities of grape pomace seeds through fungal SSF, thereby facilitating the valorization of winemaking by-products.

The effect of team cultural tightness and transformational leadership on employee creative behavior: A cross-level moderated mediation model.

In recent years, "cultural tightness-looseness" refers to the strength of social norms and tolerance for deviant behaviors perceived by individuals in society, and team cultural tightness applied at the organizational level represents the strength of team norms. According to the cultural tightness-looseness theory, this study explores the influence of team cultural tightness on employee creative behavior, as well as the moderating role of transformational leadership and the mediating role of work engagement. A questionnaire method was used to survey five enterprise organizations in China through three stages, and 288 paired questionnaires were finally obtained from leaders and employees. The results found that team cultural tightness has a significant negative effect on employee creative behavior through the mediating effect of work engagement. In other words, the more the team culture in the workplace tends to be loose, the more it stimulates employee work engagement, which in turn promotes employee creative behavior. In addition, transformational leadership played a significant positive moderating role in the model. Under the influence of transformational leadership style, the negative impact of team cultural tightness on employee work engagement and creative behavior can be mitigated. The study enriches the understanding of the influence and action mechanism of the cultural tightness-looseness theory on employee creative behavior. In future research, the action mechanism of team cultural tightness interacting with different leadership styles to influence employee creative behavior will be explored more deeply.

HIV-related stigma acting as predictors of unemployment of people living with HIV/AIDS.

Obtaining employment is an important part of recovery for many people living with HIV/AIDS (PLHA). However, this population often faces barriers in their attempt to reenter the workplace. One potential barrier lies in the decision-making of employers. Little is known about what influences employers' decision to hire PLHA. The current paper addresses this gap with findings from 156 quantitative interviews with employers across Chicago, Beijing, and Hong Kong regarding the hiring of people with HIV/AIDS. Hierarchical regression analysis showed that both fear of contagion and perceived incompetence are important factors in employers' decision to interview even after controlling for variables related to the employers' business size, their education level, and the provision of health benefits. These two variables accounted for 42% of the variance in employers' decision to interview. Implications of these findings are considered for better understanding of HIV-related employment stigma and further intervention for employing PLHA.

Sino-American employer perspective about behavioral-driven health conditions: predictive analyses.

OBJECTIVE: A large-scale quantitative study was conducted by stratified representative samples from Chicago (prototype of the United States; N = 293), Beijing (prototype of urbanized China; N = 302), and Hong Kong (prototype of East-meets-West culture; N = 284) to explore factors that might lead to their stigmatizing attitudes towards hiring individuals with (mental illness, alcohol abuse, drug abuse, and HIV/AIDS) and without (bone cancer) behavioral-driven health conditions. METHODS: Consented employers completed the Employer Survey pertaining to their attitudes towards specific health conditions, previous hiring experiences, resources, assets of applicants, and hiring concerns. RESULTS: The findings suggested that employers in Hong Kong and Beijing were more willing to hire individuals with alcohol abuse, whereas employers in Chicago were more willing to hire those with HIV/AIDS or bone cancer. Logistic regression suggested that the type of health conditions, assets of applicants, and perceived level of dangerousness of applicants were significant predictors that contributed to employers' hiring preference. CONCLUSION: Employers express different hiring preference towards individuals with or without behavioral-driven health conditions. Their hiring preference towards specific type of health conditions is discussed.

Mechanisms of Organizational Cultural Tightness on Work Engagement during the COVID-19 Pandemic: The Moderating Role of Transformational Leadership.

Cultural tightness-looseness, one of the cultural dimensions that play an essential role in organizational development, is changing the perception of psychology and behavior in organizations. This study conducted a paired questionnaire survey of leaders and their employees from five Chinese companies over three periods during the COVID-19 pandemic. The results found that organizational cultural tightness was more influenced by transformational leadership. Different from previous findings, in the context of the Chinese epidemic, organizational cultural tightness positively predicted employees' work engagement with the moderating effect of transformational leadership. Team-member exchange also mediated employees' work engagement, which had a facilitative effect on employees' work engagement. In future research, the contingent effects of other leadership styles and organizational cultural tightness will be explored to reveal the different mechanisms of action on employees' work engagement.