Metagenomic Insights into the Microbial Composition and Functional Potential of Cocoa (Theobroma Cacao L.) During Fermentation and Drying in Colombia.

Shotgun metagenomics is an approach increasingly applied to investigate microbial succession and functional potential in complex fermented food systems, including cocoa bean fermentation. In this study, we used Illumina-based shotgun metagenomic sequencing to characterize microbial community dynamics and metabolic potential across two post-harvest cocoa processing routes (R1 and R2) in Boyacá, Colombia, encompassing both fermentation and drying stages. Cocoa beans were sampled at defined time points during fermentation and subsequent natural drying, and non-host metagenomic reads were subjected to taxonomic classification and functional annotation to assess fungi, bacteria, and viruses. A clear multi-ecological succession was observed throughout post-harvest processing. Fungal communities shifted from a yeast-dominated profile, mainly Saccharomyces and Pichia during fermentation, to the emergence of the filamentous fungus Aspergillus during drying. Bacterial populations transitioned from diverse Enterobacteriaceae in early fermentation to a near-complete dominance of Acetobacter, which persisted throughout the drying phase. Viral communities also displayed structured successional patterns, with Lambdavirus and Punavirus prevalent in early fermentation, followed by Spbetavirus, Lafunavirus, and Pemunavirus during later stages and drying. Functional analyses revealed high metabolic potential for carbohydrate, energy, and amino acid metabolism during early fermentation, followed by a marked reduction in later stages, indicating a metabolic slowdown. Core metabolic functions were retained during drying at substantially lower activity levels. This integrated metagenomic analysis links microbial structure to functional potential and provides a scientific basis for optimizing starter cultures and post-harvest processing strategies to enhance cocoa quality and safety.