Metagenomic and metabolomic analyses of fecal samples from civet-digested coffee in Vietnam

<h4>Background</h4>Civet-digested coffee originates from the feces of civets that consume coffee cherries, where microbial fermentation in the gastrointestinal tract imparts distinctive flavor attributes, thereby enhancing its global reputation and market value. Gut microbiota is considered important drivers of coffee-bean fermentation, potentially shaping the unique and region-specific flavor characteristics of civet-digested coffee. To address this context, the present study integrated metagenomic and metabolomic analyses to compare the gut microbiota and secondary metabolites involved in coffee-bean fermentation inside Vietnamese civets.<h4>Methods</h4>Fecal samples were collected under two dietary conditions: a standardized one containing 20% protein, 6% fiber, and 0.4-1.5% lysine, and the same diet supplemented with coffee cherries. Metagenomic 16S rRNA sequencing and untargeted ultra-performance liquid chromatography quadrupole time-of-flight (UPLC-QTOF) revealed clear differences between the two groups.<h4>Results</h4>Integrated metagenomic and metabolomic analyses revealed clear distinctions between the two groups. Civets on the coffee-cherry diet exhibited higher microbial diversity at the family and genus levels. Specifically, among 31 classified bacterial genera showing a trend toward significant differences in abundance, <i>Enterococcus</i> and <i>Escherichia/Shigella</i> decreased, whereas <i>Gluconobacter</i>, and <i>Pseudomonas</i> increased following the diet shift. Metabolomic profiling identified 46 metabolites across both ionization modes, and strong correlations were observed between microbial genera and metabolite profiles. Specifically, 6-hydroxyangolensic acid methyl ester, 4-aminobenzoic acid and caffeine were more abundant in civets on a coffee-cherry diet, meanwhile the other nine metabolites were more prevalent in the normal diet. Overall, the findings demonstrate that civet gut microbiota and metabolic output were highly responsive to dietary inputs, and that coffee cherries promoted a unique fermentation environment. This represents the first integrative metagenomic and metabolomic study of civets consuming coffee in Vietnam, providing valuable insights into microbial contributions to coffee fermentation.