Abstract

Cacao fermentation is one of the fundamental processes for generating chocolate flavor and aroma. Fermentation process is catalyzed by environmental microbes that colonize and degrade the cacao pulp producing a wide range of metabolic end-products. Currently, there is a large amount of information related to the role of yeast, lactic acid bacteria, and acetic acid bacteria in cacao fermentation which contrasts with the limited information available for the bacteria belonging to the Enterobacterales order. These microbes might be capable of metabolizing the cacao pulp-bean mass. Elucidating the function of the members from this order is fundamental to improving the process. Here, we described the first approach to determine the relevance of these families during the fermentation process in several farms around Colombia, where we coupled the use of gene marker (16S rRNA) community profiling with metagenomics and gene expression analysis. Our metataxonomic and metagenomic results are in accordance with previously described microbial succession in cacao fermentation. From these we reconstructed 11 bacterial genomes from metagenomes, and reconstructed some of their metabolic capabilities, and found that several groups have the enzymatic capacity for pectin degradation. We showed that genes related to the pectin depolymerization of Tatumella species have a higher gene expression activity during the first 24 hours than previously anticipated. The expression of these genes during fermentation was assessed by RT-qPCR, using cacao bean mass fermented for 24 hours. In summary, our results show that bacteria from the Erwinaceae family have an essential role in the initial steps of fermentation.