Search Results (11,200)

Nanobodies are characterized by their small size, high specificity, and strong affinity, making them promising antiviral agents. In this study, a dual-plasmid yeast surface display (YSD) system based on the Saccharomyces cerevisiae a-agglutinin (Aga1p-Aga2p) platform was evaluated for the functional presentation of H5-specific nanobody. To investigate the influence of fusion design on display performance, enhanced green fluorescent protein (EGFP) was fused to Aga2p in two different orientations. Both configurations enabled successful surface display, while the EGFP-AGA2 orientation showed significantly higher display efficiency than AGA2-EGFP (p < 0.001). This optimized configuration was subsequently used to display Nb10, a broadly neutralizing nanobody targeting the hemagglutinin (HA) protein of H5 influenza viruses. Indirect ELISA, immunofluorescence, and confocal microscopy confirmed successful surface localization of Nb10, while flow cytometry revealed 22.10% positive cells compared with 0.30% in the negative control (p < 0.001). In hemagglutination inhibition (HI) assays, the YSD-Nb10 strain exhibited an HI titer of 3log2, whereas no detectable HI activity was observed in the control strain. Collectively, these results demonstrate the feasibility of displaying a functional H5-specific nanobody using a dual-plasmid YSD system and highlight the importance of fusion orientation for efficient surface presentation, providing preliminary practical guidance for optimization of YSD applications. Full article

Background/Objectives: Mild cognitive impairment (MCI) may precede dementia, and safe nutritional strategies able to support cognitive function are of clinical interest. Dietary nucleotides may contribute to membrane phospholipid synthesis, synaptic function, and neuroprotective pathways; however, clinical evidence in older adults with MCI remains limited. This randomized placebo-controlled trial evaluated the efficacy and tolerability of nucleotide-rich yeast extracts from Kluyveromyces fragilis and Saccharomyces cerevisiae. Methods: Seventy-two participants (mean age 73.5 ± 7.7 years; range 60–85) were randomly assigned (1:1:1) to receive K. fragilis extract, S. cerevisiae extract, or placebo once daily for 180 days. Cognitive outcomes were assessed using the Montreal Cognitive Assessment (MoCA) and Mini-Mental State Examination (MMSE) at baseline (T0), 90 days (T1), and 180 days (T2); quality of life was assessed using the SF-12 questionnaire at T0 and T2. Treatment effects were analyzed using linear mixed-effects models adjusted for age and sex. Results: After 180 days, MoCA scores increased by 4.42 points in the K. fragilis group and 3.92 points in the S. cerevisiae group, compared with 0.58 points in the placebo group (time × treatment p < 0.001; T0–T2 within-group p < 0.001 for both active groups and p = 0.14 for placebo). MMSE scores increased by 1.62 and 3.11 points in the K. fragilis and S. cerevisiae groups, respectively, compared with 0.25 points in the placebo group (time × treatment p < 0.001; T0–T2 within-group p < 0.001 for both active groups and p = 0.57 for placebo). The SF-12 mental component score increased by 7.50 and 9.16 points in the two active groups, respectively (time × treatment p = 0.022; T0–T2 p = 0.0013 and p < 0.001, respectively), while physical quality-of-life scores did not change significantly (PCS time × treatment p = 0.11). No adverse events were reported. Conclusions: Nucleotide-rich K. fragilis and S. cerevisiae yeast extracts were well tolerated and were associated with improved cognitive scores over six months in older adults with MCI. Larger multicenter trials are needed to confirm these findings. Full article

In Italy, the apple cider market is experiencing significant growth, driven by numerous small-scale artisanal producers who combine local apple varieties with traditional processes to offer complex, and diverse products. However, artisanal production based on spontaneous fermentations often encounters challenges in qualitative reproducibility, particularly related to sensory issues (stability across different vintages and high turbidity of the product). In this context, a methodology has been developed to optimize the technological process of cider production at Contrada Contro in the Monti Sibillini (MC), in Marche region, Italy. The research focused on the isolation and selection of indigenous yeasts from frozen must prepared in the 2023 vintage. Following isolation and preliminary characterization, the indigenous yeasts were used to referment the still cider, followed by 7 months of bottle aging, and a second sampling point was conducted after 14 months of aging on lees. Destructive analyses using HPLC-DAD and GC-MS were conducted to evaluate polyphenols and volatile compounds, while non-destructive analyses with a 12-quartz microbalance electronic nose and near infrared (NIR) spectroscopy allowed for a quicker assessment of production techniques. Chromatographic analysis results showed that the sensory quality of refermented products was strongly influenced by the composition of the yeast strains used. All fermentations inoculated with selected yeasts exhibited lower turbidity compared to spontaneous fermentation. These findings indicate that the selection of indigenous yeasts for cider refermentation enables the production of a high-quality product, enriched with beneficial compounds and characterized by a strong terroir identity, underscoring the importance of microbiological terroir. Full article

Lemon processing by-products are rich in flavonoids, limonoids, and phenolic acids, but their direct utilization is limited by glycoside-dominant flavonoid profiles, bitterness-associated limonoids, and insufficiently defined valorization strategies. This study compared eight food-relevant microorganisms, including lactic acid bacteria, Bacillus, yeast, and filamentous fungi, using a common aerobic submerged fermentation framework for lemon by-products. Rather than evaluating fermentation as a single uniform process, the study aimed to determine whether different microbial groups could redirect the same substrate toward distinct functional remodeling profiles. Targeted HPLC analysis of flavonoids, limonoids, and phenolic acids, together with DPPH and ABTS radical-scavenging assays, revealed clear strain-dependent differences in metabolite remodeling and antioxidant outcomes. L. plantarum showed the most consistent antioxidant enhancement profile, characterized by increased hesperetin and phenolic acid responses together with low DPPH and ABTS IC₅₀ values. L. pentosus promoted flavonoid remodeling but showed a more timing-sensitive antioxidant response. S. cerevisiae tended to preserve glycosylated flavonoids and showed a release-oriented phenolic acid profile with strong early ABTS activity. R. stolonifer exhibited the most pronounced limonoid remodeling, including marked limonin reduction and obacunone accumulation, suggesting potential relevance for bitterness-oriented applications. These findings demonstrate that different microorganisms can be functionally classified according to their dominant remodeling tendencies, including antioxidant enhancement, flavonoid conversion, glycosylated flavonoid preservation, phenolic acid release, and limonoid-associated debittering. This functional classification provides a practical basis for selecting microorganisms according to the intended application of lemon by-product valorization. Full article

Low temperature constitutes a critical abiotic stress that significantly impairs plant growth and development, particularly for species in cold regions. In Northeast China, the persistently low winter temperatures over an extended period pose significant challenges to the survival of chrysanthemums. This study employed the ground cover plant ‘Yingjie’ as the experimental material and cloned CmMYBS3. The CmMYBS3 protein lacks transcriptional activity and is localized exclusively in the nucleus. Under low-temperature treatment, the activities of SOD, CAT, and POD were significantly lower in chrysanthemums overexpressing CmMYBS3 than in the wild-type line. Additionally, the MDA content in the CmMYBS3 overexpression lines was higher than in the wild-type lines. To elucidate the mechanism by which CmMYBS3 regulates the response to low temperature, we conducted transcriptome sequencing analysis and identified a total of 5425 differentially expressed genes, comprising 2646 upregulated genes and 2779 downregulated genes. The GO analysis reveals that the primary enrichment occurs in the “biological process”, “cellular component”, and “molecular function”. The KEGG enrichment analysis identified significant alterations in several pathways associated with plant growth and development, as well as stress responses. Through yeast single-hybrid analysis, it was demonstrated that CmMYBS3 specifically binds to the promoter region of CmDREB1 and inhibiting the expression of the CmDREB1. This study demonstrates that CmMYBS3 reduces the cold tolerance of ground cover chrysanthemums by suppressing the expression of the CmDREB1 gene, providing an important theoretical basis for the breeding of cold-tolerant ground cover chrysanthemum varieties. Full article

This paper presents the results of testing a copper bioleaching technology applied to two types of ore sampled from different sections of deposits within one of the deposits in the Balkhash region. Preliminary microbiological studies of microorganisms present in mineral raw material samples from the deposit revealed that, under conditions favorable for the growth of iron- and sulfur-oxidizing bacteria, active proliferation of yeast-like fungi was also observed, along with a bacterial culture identified as Skermanella aerolata. Preliminary experiments demonstrated that the effect of the identified bacterial culture, in association with Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans, positively influences oxidative processes involved in the decomposition of sulfur- and iron-containing minerals. The complete consortium of endemic microorganisms used in bioleaching experiments exhibited the highest efficiency compared to both individual cultures and the conventional sulfuric acid leaching method. The effect of biological oxidation on a simple-composition ore sample resulted in a 5.4% increase in copper recovery, while the efficiency of sulfuric acid consumption improved by nearly 40%. The use of bacterial oxidation for a low-grade, high-acid-consuming ore sample showed comparable copper recovery; however, sulfuric acid consumption was reduced by a factor of 2.5. Full article

Cadmium (Cd) toxicity poses a significant threat to crop production and food safety. Although proline is known to enhance plant tolerance to Cd, the molecular mechanisms regulating Cd detoxification through proline accumulation remain unclear. This study identifies the proline transporter TaPROT2 as a crucial positive regulator of Cd tolerance in wheat. We demonstrate that overexpression of TaPROT2 directly promotes proline accumulation in transgenic wheat while simultaneously activating the antioxidant enzyme system, thereby reducing both Cd accumulation and translocation. Using electrophoretic mobility shift assays (EMSA), yeast one-hybrid (Y1H) assays, and luciferase reporter assays, we confirmed that TaERF3 directly binds to the GCC-box element in the TaPROT2 promoter, thereby activating its transcription. Furthermore, overexpression of TaERF3 enhances the expression of TaPROT2, leading to increased proline accumulation and decreased Cd content. In summary, our study reveals a novel TaERF3-TaPROT2 module that promotes proline accumulation, reduces Cd accumulation, and enhances Cd tolerance, providing a promising target for breeding low-Cd wheat. Full article

Kombucha is traditionally produced by fermenting Camellia sinensis tea and sugar in a consortium of microorganisms called SCOBY (Symbiotic Culture Of Bacteria and Yeasts). Short- and medium-chain fatty acids and other organic acids in K are mainly produced by acetic acid bacteria, which contribute to the typical K taste. Coffee is one of the most widely consumed beverages in the world and one of the most traded commodities globally. Harvesting during coffee production generates tons of byproducts generally considered of low value, including cascara (CC), composed of dried pulp and skin, and leaves (CL). To date, few studies have investigated the production of short- and medium-chain fatty acids and monosaccharide’s profile during traditional kombucha fermentation, and their composition in kombuchas prepared from substrates other than C. sinensis is even scarcer. This study followed the changes in sugars and the production of short- and medium-chain fatty acids during K fermentation of black tea (BT), CC, and CL and associated their concentrations with physicochemical parameters (total soluble solids (TSS), pH, and titratable acidity (TA)) and the perceived acidity of the beverages evaluated by a trained panel and untrained consumers. BT K, a SCOBY, and 10% sucrose were added to infusions of arabica CC, CL, or BT. The mixture was fermented for 0, 3, 6, and 9 days. Organic acids were analyzed by GC-MS; sucrose and monosaccharides were analyzed by HPLC-RID. The Rate All That Apply (RATA) test was used for sensory analysis. Results were treated by ANOVA–Fisher and Pearson correlation tests with significance at p < 0.05. Glucose, fructose, arabinose, xylose, cellobiose and glycerol were identified in the infusions. On average, sucrose concentration decreased by 28% up to day 9, considering all K samples, accompanied by TSS decrease. Eight organic acids were semi-quantified, with acetic being the major acid in all beverages (8.4 to 1971 mg L⁻¹) and isovaleric being the lead minor acid (0.7 to 17.7 mg L⁻¹). Additional acids identified were: butanoic, 2-methylpropanoic, pentanoic, 3-methylpentanoic, hexanoic, and octanoic acids. TA values and sourness perceived by consumer assessors increased generally, even though in CC Ks, the acid concentration decreased by day 9. TA, sourness, and sparkling and fizzy mouthfeel correlated positively in all Ks. In general, although the total acid concentration was mainly higher on days 3 or 6, CO₂ formation, among other organic acids, probably increased TA and sourness on day 9. Although it is generally accepted that pH and organic acid concentrations are directly associated with sour taste, it is not possible to accurately predict and modify sour taste intensity in kombucha based only on these parameters, given that other factors, such as the production of CO_2, the existence of buffer systems, and the presence of sugars and other soluble solids, will probably affect the perceived acidity and sourness. Full article

Non-Saccharomyces yeasts (NSY) are increasingly investigated as biotechnological tools to diversify wine profiles and modulate fermentation outcomes. This study evaluated the enological behavior of two Chilean isolates, Starmerella bacillaris (SB) and Hanseniaspora uvarum (HU), in Sauvignon Blanc must from the Casablanca Valley under monoculture and sequential inoculation (NSY → Saccharomyces cerevisiae) at laboratory (500 mL) and microvinification (10 L) scales. In synthetic medium (150 g/L sugars), SB and HU showed incomplete sugar consumption, producing 4.25% and 8.50% v/v ethanol, respectively, compared with 9.16% v/v for S. cerevisiae. In laboratory-scale fermentation in real must, both strains completed fermentation in monoculture, with moderate reductions in ethanol production relative to the control. At the microvinification scale, monocultures yielded lower ethanol concentrations (11.90–12.50% v/v) than S. cerevisiae (13.50% v/v), whereas sequential fermentations converged toward control values. NSY treatments showed higher relative abundances of medium-chain ethyl esters associated with fruity and floral sensory attributes while maintaining acetic acid concentrations ≤ 0.50 g/L. These findings indicate that the effects of SB and HU depended primarily on fermentation strategy and process scale under the evaluated conditions. Full article

Leptolegnia chapmanii is an oomycete pathogen of mosquito larvae. We investigated whether nutritional factors promoting cyst germination in vitro are associated with early infection events and instar-specific susceptibility in Aedes aegypti. Cysts of the Brazilian isolate ARSEF 12829 germinated rapidly in soybean seed extract, sunflower seed extract and minimal medium supplemented with yeast extract, whereas basal minimal medium did not promote germination. In sunflower seed extract, germination increased significantly with incubation time; in minimal medium, germination at 24 h was much higher with ≥0.2% yeast extract than with 0.1%. In third-instar larvae, a few cysts attached to the cuticle during the first 30–60 min, with no external germ tubes observed. At 3 h, melanized hyphal structures were detected in the midgut, and histological sections showed germinated and ungerminated cysts in the endoperitrophic space, with hyphae crossing the peritrophic matrix and midgut epithelium toward the hemocoel. Mortality increased with cyst concentration and exposure time and decreased with larval instar. At 3.3 × 10³ cysts/mL, final mortality reached 100% in L1–L3 and 91.2% in L4 larvae. These results link rapid cyst germination with early midgut invasion and high larvicidal activity. Full article

Expression of the mevalonate (MVA) pathway in yeast mitochondria is deployed at commercial scale for production of squalene, because mitochondria contain abundant acetyl-CoA, the starting molecule of the MVA pathway. However, it is still unknown whether this strategy is effective in boosting the post-squalene pathway. Here the potential of this strategy is explored for production of the post-squalene chemical cholesterol, a precursor of many valuable steroidal drugs. A cholesterol-producing yeast strain, named CEN-Cho, was constructed by expressing the biosynthetic genes leading to cholesterol, accompanied by the augmentation of the cytosolic MVA pathway. The CEN-Cho strain produced 60.17 ng/mg fresh weight (FW) of squalene and 121.75 ng/mg FW of cholesterol in shake flask cultivation. When the complete MVA pathway was introduced into the mitochondria of CEN-Cho, yielding CEN-Cho-mMVA, the squalene level was increased to 325.24 ng/mg FW. Unexpectedly, the yield of cholesterol produced by CEN-Cho-mMVA was decreased to 1.5 ng/mg FW, demonstrating significant suppression on the post-squalene pathway due to the mitochondrial engineering. Comparative transcriptomic and proteomic analyses of the engineered cells provide insights into the metabolic and regulatory bottlenecks underlying this inhibition. This work reveals that the introduction of the MVA pathway into mitochondria generally inhibits the post-squalene pathway in yeast. Full article

Pear fruits host diverse microbial communities that influence postharvest quality, spontaneous fermentation, and susceptibility to microbial contamination. This study characterizes the fungal communities associated with naturally fallen overripe pears (Pyrus communis) using ITS2 amplicon sequencing combined with culture-dependent approaches. The fungal community exhibited low diversity and was dominated by Ascomycota (99%), primarily Saccharomycetes (91.8%), with Hanseniaspora, Aureobasidium, and Microcyclospora representing more than 90% of the total microbial community. Culture-dependent isolation confirmed Hanseniaspora uvarum as the dominant yeast species (~89%), followed by Metschnikowia spp. and Pichia spp. Pairwise co-culture assays, quantified using the Relative Interaction Index, demonstrated predominantly competitive interactions, with fast-growing H. uvarum exerting suppressive effects on slower-growing species. Among the isolated yeasts, Metschnikowia fructicola exhibited antibacterial activity against all tested bacteria Staphylococcus aureus, Listeria innocua and Salmonella typhimurium. The strongest antibacterial activity was exerted against the foodborne pathogen S. aureus. In a pear juice model system, co-cultivation with M. fructicola resulted in the elimination of S. aureus within four days, while yeast viability was maintained. These findings observe the fermentative yeasts distributed in overripe pears and demonstrate the potential of M. fructicola to inhibit bacterial growth under controlled conditions. The results provide a preliminary basis for further studies on fungal succession, yeast interactions, and the biocontrol potential of pear-associated yeasts. For broader ecological conclusions, larger-scale studies across locations, seasons, cultivars, and decay stages are required. Full article

Selenium-enriched yeast is an important organic selenium source for improving the nutritional value and physiological function of aquatic products. However, whether ploidy affects the molecular responses of Pacific oysters (Crassostrea gigas) under selenium supplementation conditions remains unclear. In this study, diploid, triploid, and tetraploid Pacific oysters were fed a selenium-enriched diet for 10 days. Hepatopancreas tissues were collected for antioxidant enzyme assays and transcriptomic analysis. The results showed that triploid oysters exhibited higher superoxide dismutase, glutathione peroxidase, and catalase activities, together with a relatively lower malondialdehyde level, suggesting stronger antioxidant defense under selenium supplementation conditions. Transcriptomic analysis revealed substantial numbers of differentially expressed genes across all groups compared, indicating pronounced ploidy-dependent transcriptional responses. Functional enrichment analysis showed that these genes were mainly associated with metabolic processes, oxidative stress responses, membrane components, endocrine-related pathways, lipid metabolism, and signal transduction. Several redox-related genes, including glutathione S-transferase family genes, CHAC1, superoxide dismutase family genes, and NRX1, showed ploidy-dependent expression patterns. In particular, triploid oysters appeared to rely more on enzymatic antioxidant defense and glutathione turnover, whereas tetraploid oysters showed a stronger tendency toward redox signal integration and homeostatic regulation. These findings provide new insights into ploidy-dependent selenium responses in Pacific oysters and offer a theoretical basis for the development and utilization of selenium-enriched oyster products. Full article

This study aimed to analyze the effect of various phenolic acids introduced into pectin films on their ability to inhibit microorganisms. The antimicrobial activity of six phenolic acids—gallic, protocatechuic, caffeic, sinapic, coumaric, and ferulic acids—was verified against Bacillus subtilis bacteria. No inhibitory effect was observed when the acids were introduced into the substrates with the films, as the polysaccharide films served as a breeding ground for microorganisms. Bacterial growth was inhibited when pure acid was introduced to the substrate. Gallic and caffeic acid, at concentrations of 50 and 75 mM/dm³, respectively, completely inhibited bacterial growth. However, studies on pears have shown that such concentrations of phenolic acids are unsuitable for fruit coatings, as they lead to cloudiness and impaired visual appeal. Consequently, the lowest effective concentration was applied to fruit, reducing the total bacterial count from 2.59 ± 0.04 to 1.88 ± 0.22 log CFU/mL and mold and yeast counts from 2.11 ± 0.09 to 1.63 ± 0.10 log CFU/mL. The coating produced with the lowest tested concentration of gallic acid reduced the pears’ respiration rate. The amount of CO₂ released by coated fruit was approximately 4 mg/kg·h lower than that of uncoated pears, and the level of ethylene released was approximately 6 ppm lower. The addition of gallic acid at a concentration of 15 mM/dm³ to the coating reduced the growth of bacteria, yeasts, and molds. After 12 days of pear storage, the number of microorganisms in coated fruit was approximately 0.71 log CFU/mL lower for bacterial cells and 0.48 log CFU/mL lower for yeasts and molds. Full article

The increasing volume of discarded apples generated by commercial grading standards and postharvest losses represents both an environmental burden and an opportunity for sustainable valorization. Despite growing interest in circular economy strategies in the fruit-processing sector, a comprehensive review of the technological, microbiological, and nutritional factors influencing cider production from discarded apples remains limited. To address this gap, this review discusses key aspects of cider production from discarded apples, focusing on raw material characterization, nutrient management, yeast strategies, and fermentation technologies. The physicochemical and microbiological properties of discarded apples are examined, including soluble solids, acidity, phenolic composition, and microbial spoilage risks. Special attention is given to nutrient optimization, particularly yeast assimilable nitrogen (YAN), vitamins, and minerals, as deficiencies may cause sluggish fermentation and adversely affect volatile compound formation and stability. This review evaluates yeast selection, comparing Saccharomyces cerevisiae with non-Saccharomyces yeasts and mixed fermentations, highlighting their effects on chemical composition, aroma, and sensory quality. Innovative approaches such as yeast immobilization and repeated-batch fermentation are reviewed as tools to improve process performance. Key technical challenges, including variability in raw material quality, nutrient supplementation needs, contamination risks, and process scalability, are discussed alongside opportunities for valorization of cider pomace within a circular economy framework. Full article