ABSTRACT
The microbiome in fermentation has direct impacts on the quality of fermented foods and is of great scientific and commercial interest. Despite considerable effort to explain the microbial metabolism associated with food fermentation, the role of the microbiome in pu-erh tea fermentation remains unknown. Here, we applied integrated meta-omics approaches to characterize the microbiome in two repeated fermentations of pu-erh tea. Metabarcoding analysis of bacterial 16S rRNA genes showed a decrease in the proportion of Proteobacteria and an increase in the abundance of Firmicutes during fermentation. Metabarcoding analysis of fungal internal transcribed spacer (ITS) sequence demonstrated that Rasamsonia, Thermomyces, and Aspergillus were dominant at the intermediate stage, whereas Aspergillus was dominant at other stages in fermentation. Metaproteomics analysis assigned primary microbial metabolic activity to metabolism and identified microbial carbohydrate-active enzymes involved in the degradation of polysaccharides including cellulose, xylan, xyloglucan, pectin, starch, lignin, galactomannan, and chitin. Metabolomics and high-performance liquid chromatography analysis revealed that levels of phenolic compounds, including gallates, decreased whereas contents of gallic acid and ellagic acid significantly increased after fermentation (P < 0.05). The changes in levels of gallates and gallic acid were associated with the hydrolysis of tannase. Glycoside hydrolases, phenol 2-monooxygenase, salicylaldehyde dehydrogenase, salicylate 1-monooxygenase, catechol O-methyltransferase, catechol dioxygenase, and quercetin 2,3-dioxygenases were hypothesized to be related to oxidation, conversion, or degradation of phenolic compounds. We demonstrated microbiota in fermentation and their function in the production of enzymes related to the degradation of polysaccharides, and metabolism of phenolic compounds, resulting in changes in metabolite contents and the quality of pu-erh tea.IMPORTANCE Fermented foods play important roles in diets worldwide and account for approximately one-third of all foods and beverages consumed. To date, traditional fermentation has used spontaneous fermentation. The microbiome in fermentation has direct impacts on the quality and safety of fermented foods and contributes to the preservation of traditional methods. Here, we used an integrated meta-omics approach to study the microbiome in the fermentation of pu-erh tea, which is a well-known Chinese fermented food with a special flavor and healthful benefits. This study advanced the knowledge of microbiota, metabolites, and enzymes in the fermentation of pu-erh tea. These novel insights shed light onto the complex microbiome in pu-erh fermentation and highlight the power of integrated meta-omics approaches in understanding the microbiome in food fermentation ecosystems.
ABSTRACT
A high frequency of death-associated protein kinase (DAPK) promoter hypermethylation has been noted in B-cell malignancies, head and neck cancers, and other solid tumors, and it has been used as a tumor marker in molecular detection strategies. Low levels of DAPK promoter hypermethylation, ranging from 0.003 to 1.181%, were detected in peripheral blood cells from 75 of 143 (52%) normal subjects by quantitative methylation-specific PCR (Q-MSP). In 10 of 10 selected patients, MSP amplification of a portion of the DAPK promoter followed by PCR product sequencing confirmed dense hypermethylation of the CpG island in their peripheral blood cells. Q-MSP analysis of fluorescence-activated cell-sorted peripheral blood cells from three of these patients demonstrated that a significantly greater proportion of B cells (1.074-6.026%) were DAPK hypermethylated than were T cells, monocytes, or neutrophils, which were <0.06% hypermethylated. Further analysis after sorting of one subject's B cells into IgM+, IgM-, IgG+, and IgG- subpopulations demonstrated that DAPK hypermethylation was predominantly present in the IgM- compared with IgM+ B cells (3.338% versus 0.436%). DAPK promoter hypermethylation was found in IgM- B cells in normal individuals. The same hypermethylation identified in B-cell malignancies may reflect a clonal outgrowth of B cells arising from this compartment and may indicate a susceptibility to neoplastic transformation in a subset of B cells. Normal circulating lymphocytes with DAPK promoter hypermethylation may act as confounding factors in tumor detection based on DAPK hypermethylation.