Differential consumption of malic acid and fructose in apple musts by Pichia kudriavzevii strains.

AIMS: To assess the capability of Pichia kudriavzevii strains isolated from wine, cider, and natural environments in North Patagonia to produce ciders with reduced malic acid levels. METHODS AND RESULTS: Fermentation kinetics and malic acid consumption were assessed in synthetic media and in regional acidic apple musts. All P. kudriavzevii strains degraded malic acid and grew in synthetic media with malic acid as the sole carbon source. Among these strains, those isolated from cider exhibited higher fermentative capacity, mainly due to increased fructose utilization; however, a low capacity to consume sucrose present in the must was also observed for all strains. The NPCC1651 cider strain stood out for its malic acid consumption ability in high-malic acid Granny Smith apple must. Additionally, this strain produced high levels of glycerol as well as acceptable levels of acetic acid. On the other hand, Saccharomyces cerevisiae ÑIF8 reference strain isolated from Patagonian wine completely consumed reducing sugars and sucrose and showed an important capacity for malic acid consumption in apple must fermentations. CONCLUSIONS: Pichia kudriavzevii NPCC1651 strain isolated from cider evidenced interesting features for the consumption of malic acid and fructose in ciders.

Hi-C metagenomics facilitate comparative genome analysis of bacteria and yeast from spontaneous beer and cider.

Sequence-based analysis of fermented foods and beverages' microbiomes offers insights into their impact on taste and consumer health. High-throughput metagenomics provide detailed taxonomic and functional community profiling, but bacterial and yeast genome reconstruction and mobile genetic elements tracking are to be improved. We established a pipeline for exploring fermented foods microbiomes using metagenomics coupled with chromosome conformation capture (Hi-C metagenomics). The approach was applied to analyze a collection of spontaneously fermented beers and ciders (n = 12). The Hi-C reads were used to reconstruct the metagenome-assembled genomes (MAGs) of bacteria and yeasts facilitating subsequent comparative genomic analysis, assembly scaffolding and exploration of "plasmid-bacteria" links. For a subset of beverages, yeasts were isolated and characterized phenotypically. The reconstructed Hi-C MAGs primarily belonged to the Lactobacillaceae family in beers, along with Acetobacteraceae and Enterobacteriaceae in ciders, exhibiting improved quality compared to conventional metagenomic MAGs. Comparative genomic analysis of Lactobacillaceae Hi-C MAGs revealed clustering by niche and suggested genetic determinants of survival and probiotic potential. For Pediococcus damnosus, Hi-C-based networks of contigs enabled linking bacteria with plasmids. Analyzing phylogeny and accessory genes in the context of known reference genomes offered insights into the niche specialization of beer lactobacilli. The subspecies-level diversity of cider Tatumella spp. was disentangled using a Hi-C-based graph. We obtained highly complete yeast Hi-C MAGs primarily represented by Brettanomyces and Saccharomyces, with Hi-C-facilitated chromosome-level genome assembly for the former. Utilizing Hi-C metagenomics to unravel the genomic content of individual species can provide a deeper understanding of the ecological interactions within the food microbiome, aid in bioprospecting beneficial microorganisms, improving quality control and improving innovative fermented products.

Differential adaptation of the yeast Candida anglica to fermented food.

A single strain of Candida anglica, isolated from cider, is available in international yeast collections. We present here seven new strains isolated from French PDO cheeses. For one of the cheese strains, we achieved a high-quality genome assembly of 13.7 Mb with eight near-complete telomere-to-telomere chromosomes. The genomes of two additional cheese strains and of the cider strain were also assembled and annotated, resulting in a core genome of 5966 coding sequences. Phylogenetic analysis showed that the seven cheese strains clustered together, away from the cider strain. Mating-type locus analysis revealed the presence of a MATa locus in the cider strain but a MATalpha locus in all cheese strains. The presence of LINE retrotransposons at identical genome position in the cheese strains, and two different karyotypic profiles resulting from chromosomal rearrangements were observed. Together, these findings are consistent with clonal propagation of the cheese strains. Phenotypic trait variations were observed within the cheese population under stress conditions whereas the cider strain was found to have a much greater capacity for growth in all conditions tested.

Association of exposure factors and their causal relationship with oral cancer: A Mendelian randomization study.

OBJECTIVES: There is a strong association among risk factors for oral cancer (ORCA), such as smoking, alcohol consumption, fiber intake, and red meat intake. The apparent synergistic effects reported in previous observational studies may also underestimate the independent effects. Our study aims to further explore the potential etiology and causality of oral cancer. MATERIALS AND METHODS: This study used the genome-wide associations study database (GWAS) in European populations for Mendelian randomization (MR) analysis to explore exposure factors associated with ORCA and detect the genetic causality between these exposures and ORCA risk. RESULTS: Our results demonstrated that in univariate MR analysis, the five exposure factors (celery intake, average weekly beer and cider intake, spirits intake, and pork intake) were risk factors, and oily fish intake was a safety factor, but in multivariate MR analysis, pork intake had the greatest impact on oral cancer when the five food/drink intakes were simultaneously consumed. CONCLUSIONS: The causal relationship between the five exposure factors (oily fish intake, celery intake, pork intake, average weekly beer and cider intake, and spirits intake) and oral cancer was analyzed. The causal effects of pork on oral cancer may be underestimated. CLINICAL RELEVANCE: Prevention of oral cancer requires better education about lifestyle-related risk factors, and improved awareness and tools for early diagnosis. Our study provides some risk factors that cannot be ignored for the cause prevention of oral cancer, such as pork intake, and its role in oral cancer prevention and control.

Effects of Torulaspora delbrueckii and Saccharomyces cerevisiae Co-Fermentation on the Physicochemical and Flavor Compounds of Huaniu Apple Cider.

The effects of different fermentation methods utilizing Torulaspora delbrueckii 1004 and Saccharomyces cerevisiae 32169 on the physicochemical properties, organic acid content, polyphenol and flavonoid concentrations, antioxidant activity, and volatile aroma compounds of Huaniu apple cider were investigated in this study. Employing methods of single inoculation, co-inoculation, and sequential inoculation, it was found that sequential fermentation exhibited strong fermentative power in the initial stages, effectively reducing the content of soluble solids and achieving a balanced composition of malic, succinic, and citric acids while maintaining a lower titratable acidity. Sequential inoculation was observed to significantly enhance the total polyphenols and flavonoids, as well as the antioxidant capacity (p < 0.05). Specifically, in the synthesis of volatile aroma compounds, sequential inoculation significantly enhanced the richness and diversity of the Huaniu apple cider's aromas, particularly in terms of the concentration of ester compounds (p < 0.05). Principal component analysis further confirmed the superiority of sequential inoculation in terms of aroma component diversity and richness. The findings of this study suggest that sequential inoculation of fermentation with non-Saccharomyces and S. cerevisiae is an effective strategy for optimizing the flavor characteristics of Huaniu apple cider, offering valuable theoretical support and practical guidance for enhancing cider quality and fostering the development of new products.

Enhancing cider quality through co-fermentation with acid protease and esterase-producing Metschnikowia species and Saccharomyces cerevisiae.

BACKGROUND: To date, cider production has primarily relied on Saccharomyces cerevisiae. Introducing novel non-Saccharomyces yeasts can enhance the diversity of cider properties. Among these, the Metschnikowia genus stands out for its ability to produce hydrolytic enzymes that may impact the sensorial and technological properties of cider. This study focused on evaluating the impact of three Metschnikowia species - Metschnikowia koreensis (Mk), M. reukaufii (Mr), and M. pulcherrima (Mp) - which exhibit acid protease and esterase activity, on the quality enhancement of cider. RESULTS: The research findings indicate that the overall quality of cider produced through co-fermentation with these species surpassed that of cider fermented with mono-fermentation of S. cerevisiae (Sc). The cider fermented with the Sc + Mk combination exhibited the lowest levels of harsh-tasting malic acid and higher levels of softer lactic acid. Sensory array analysis also demonstrated that the Sc + Mk fermented cider exhibited high sensor response values for compounds contributing to a complex overall olfactory composition and richness. Furthermore, the Sc + Mk fermented cider exhibited the highest total quantity and variety of volatile organic compounds (VOCs). Specifically, the concentrations of phenethyl alcohol, 3-methyl-1-butanol, ethyl octanoate, and decanoic acid were notably elevated in comparison with other groups. CONCLUSION: This study illustrates that Metschnikowia species, particularly M. koreensis, show significant potential as starters for cider due to their various technological properties, including acidity modulation, aroma enhancement, and color improvement. The findings of this study provide a foundation for improving cider quality by co-fermenting S. cerevisiae with innovative starter cultures. © 2024 Society of Chemical Industry.

Impact on sales of adding a smaller serving size of beer and cider in licensed premises: an A-B-A reversal design.

BACKGROUND: Smaller serving sizes of alcoholic drinks could reduce alcohol consumption across populations thereby lowering the risk of many diseases. The effect of modifying the available range of serving sizes of beer and cider in a real-world setting has yet to be studied. The current study assessed the impact on beer and cider sales of adding a serving size of draught beer and cider (2/3 pint) that was between the current smallest (1/2 pint) and largest (1 pint) standard serving sizes. METHODS: Twenty-two licensed premises in England consented to taking part in the study. The study used an ABA reversal design, set over three 4-weekly periods, with A representing the non-intervention periods, during which standard serving sizes were served and B the intervention period when a 2/3 pint serving size of draught beer and cider was added to the existing range, along with smaller 1/2 pint and larger 1 pint serving sizes. The primary outcome was the daily volume of beer and cider sold, extracted from sales data. RESULTS: Fourteen premises started the study, of which thirteen completed it. Twelve of those did so per protocol and were included in the primary analysis. After adjusting for pre-specified covariates, the intervention did not have a significant effect on the volume of beer and cider sold per day (3.14 ml; 95%CIs -2.29 to 8.58; p = 0.257). CONCLUSIONS: In licensed premises, there was no evidence that adding a smaller serving size for draught beer and cider (2/3 pint) when the smallest (1/2 pint) and largest (1 pint) sizes were still available, affected the volume of beer and cider sold. Studies are warranted to assess the impact of removing the largest serving size. TRIAL REGISTRATION: ISRCTN: https://doi.org/10.1186/ISRCTN33169631 (08/09/2021), OSF: https://osf.io/xkgdb/ (08/09/2021).

Effects of film-forming Pichia and Candida yeasts on cider and wine as post-fermentation contaminants.

Film-forming yeasts are potential sources of defects in alcoholic beverages. The aim of this study is to assess the growth capacity of Pichia and Candida film-forming yeasts in cider and wine and the effects on their chemical composition. Cider, partially and fully fermented wine were inoculated with strains of C. californica, P. fermentans, P. kluyveri, P. kudriavzevii, P. manshurica, and P. membranifaciens to simulate a post-fermentative contamination. The former three species grew only in cider. Pichia manshurica and P. kudriavzevii displayed high viability in wine up to 13.18% (v v-1) ethanol. Significant changes in odour-active molecules from different chemical groups were observed in cider and wine in the inoculated samples, compared to the non-inoculated ones. Cider is more susceptible to contamination by all of the species tested, due to its low alcohol content, while P. membranifaciens, P. manshurica, and P. kudriavzevii are additionally potential spoilage agents of wine. This study highlights the risk of cider and wine contamination by film-forming yeasts. Their impact on aroma profiles depends on their ability to grow and their metabolism. This study contributes to an understanding of the possible physiological and metabolic mechanisms responsible for film formation and chemical changes in alcoholic beverages.

A comparative study of the effect of bacteria and yeasts communities on inoculated and spontaneously fermented apple cider.

Understanding bacteria and yeasts communities can reduce unpredictable changes of apple cider. In this study, apple juice inoculated with Saccharomyces cerevisiae WET 136 and fermented spontaneously were compared, the relationships of bacteria, yeasts, organic acids, and volatiles were analyzed. Results showed that microbial diversity affected the fermentation, organic acids and volatiles in apple ciders. In the first four spontaneous fermentation days, LAB (lactic acid bacteria) multiplied and reached 7.89 lg CFU/mL, and then triggered malolactic fermentation (MLF), leading to malic acid decreased by 3880.52 mg/L and lactic acid increased by 4787.55 mg/L. The citric, succinic and fumaric acids content was 2171.14, 701.51 and 8.06 mg/L lower than that in inoculated cider, respectively. Although the yeasts multiplied during spontaneous fermentation, it did not reach 7.50 lg CFU/mL until the 5th day, which led to a long lag period, as well as later and lower production of acetaldehyde and higher alcohols. The inoculated yeast inhibited LAB, acetic acid bacteria, Rahnella, and non-Saccharomyces. Yeasts were the key to produce citric acid, acetaldehyde and 3-methyl-1-butanol in apple cider; while bacteria were closely related to the formation of lactic acid, acetic acid and ethyl acetate. It suggested that low higher alcohols and acetaldehyde can be realized by selecting yeasts, and Leuconostoc pseudomesenteroides can work as candidate to reduce L-malic and citric acids in apple cider.

The Efficacy of Apple Cider Vinegar at Different pH Values as an Antimicrobial Agent: An In Vitro Study.

AIMS AND BACKGROUND: This study evaluates the antimicrobial activities of commercially available 5% apple cider vinegar (ACV) against Enterococcus faecalis, Streptococcus mutans, and Lactobacillus casei. Materials and methods: Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were conducted using the broth microdilution method. Sodium hypochlorite (NaOCl) of 5.25% was used as a positive control, and comparisons were also made with acetic acid (AA) as the main ingredient in ACV. The three test bacteria treated with the most effective ACV dilution were visualized under a transmission electron microscope (TEM) for structural changes. RESULTS: Minimal inhibitory concentration was determined at 0.625% of the concentration of ACV against S. mutans and E. faecalis and 1.25% of the concentration of ACV against L. casei with two-fold serial dilutions. A concentration of 5 × 10-1% with 10-fold serial dilutions was found to be the MIC value for all three bacteria. No significant differences were found when compared with the positive control (NaOCl) (p = 0.182, p = 0.171, and p = 0.234), respectively, for two-fold serial dilutions and (p = 1.000, p = 0.658, and p = 0.110), respectively for 10-fold serial dilutions. MBC was observed to be 5% ACV for both E. faecalis and S. mutans. However, positive microbial growth was observed on the agar plate when cultured with L. casei. An independent sample t-test showed no significant differences (p > 0.05) in the antimicrobial activities between 5% ACV and 5% pure AA. TEM revealed cell wall and cytoplasmic membrane disruptions on all three bacteria at MIC value. CONCLUSION: Apple cider vinegar has antimicrobial activities against Enterococcus faecalis, Streptococcus mutans, and Lactobacillus casei at their respective MIC values. CLINICAL SIGNIFICANCE: Apple cider vinegar can be an alternative antimicrobial dental pulp disinfectant to sodium hypochlorite. Apple cider vinegar can be used safely, especially in children's dental pulp therapy and deep caries management, when adequate tooth isolation is not readily achievable. Thus, adverse reactions commonly associated with other frequently used chemical disinfectants can be avoided.