Molecular identification of lactic acid bacteria involved in Ghanaian cocoa bean fermentation

Cocoa is a vital cash crop in Ghana, contributing approximately 15% of the country’s foreign exchange earnings and employing over 800,000 farmers. Fermentation significantly influences cocoa quality and flavour, yet controlling this process remains a challenge. This study investigated lactic acid bacteria (LAB) involved in Ghanaian cocoa fermentation by analyzing samples from the Eastern and Western North Regions. A combination of culture-dependent methods, 16S rRNA gene sequencing (Sanger platform), and phylogenetic analysis was used to identify and characterize LAB diversity. Bioinformatics tools, including BLASTN (NCBI) and R software (v4.4.0), were employed for taxonomy assignment and visualization, while phylogenetic analysis was performed using PhyML (v3.0) and iTOL. Three LAB species were identified: Limosilactobacillus fermentum , Lactiplantibacillus plantarum , and Lentilactobacillus hilgardii . L. fermentum dominated the fermentation process, forming two distinct phylogenetic clusters, suggesting genetic diversity. Temperature and pH analysis revealed similar trends across regions, with peak fermentation temperatures (47.1°C and 45.4°C) coinciding with the highest pH values (5.19 and 5.5) at 72 hours. Notably, L. plantarum was isolated at 72 hours, a stage characterized by a declining pulp pH (4.9) and a heap temperature of 45.4°C, suggesting its adaptation to intermediate fermentation conditions when acidification intensifies, and microbial succession progresses. Statistical analysis showed no significant regional differences in temperature (p > 0.05), but fermentation time significantly influenced both temperature and pH dynamics (p < 0.05). These findings enhance the understanding of LAB and physicochemical dynamics in cocoa fermentation, offering insights for optimizing fermentation processes to improve cocoa quality and chocolate flavour.