Abstract Description

Naturally fermented foods, especially milk-derived products in some African regions, are considered a staple food, serving as an excellent source of protein and vitamins. This has gained notable traction as potential functional foods with health properties due to its inherent probiotic populations. Traditional African fermented milks (Amasi and Madila), though widely popular amongst local communities, remain largely unexplored and possess microbes capable of surviving the harsh conditions of the human GIT. Therefore, these fermented milks may serve as a matrix for novel, strain-specific, probiotic microbes. However, the complex microbial interactions and unique regional processing practices warrant in-depth analysis to uncover their full functional potential. Hence, this study screens for probiotic strains from locally fermented Amasi and Madila (in vitro), whilst simultaneously exploring the milk microbiome (shotgun metagenomics). Findings revealed that 54% of isolates survived simulated GIT conditions, with isolate AV3-9 showing the highest tolerance (97.8%). Subsequent screening identified the key functional strains: Lactiplantibacillus plantarum MV2-2 (antibiotic susceptibility), Enterococcus faecium AV2-9 (cholesterol reduction), Saccharomyces cerevisiae MV2-1 (GABA production), and Lactiplantibacillus plantarum AV3-9 (best overall performance). Metagenomics (alpha/beta diversity) revealed vendor as the most significant factor shaping taxonomy, pathways, and genes (p < 0.001). Lactococcus cremoris dominated across samples, while selected probiotics (in vitro) came from rarer taxa, emphasising the need for strain-specific screening. Probiotic-linked genes that were dominant were those encoding acid tolerance. This study thus highlights multifunctional probiotic traits (in vitro) and probiogenomic insights, positioning artisanal fermented milk as a promising source of novel probiotics for nutrition, health, and sustainable food security.