Probiotic Apilactobacillus Kunkeei EABW06 from Beeswax Microbiota: A Candidate for Functional Dairy Foods

Apilactobacillus kunkeei, a fructophilic lactic acid bacterium (FLAB) associated with the honeybee microbiota, has recently gained attention as an unconventional probiotic source. This study evaluated the probiotic, safety, and technological properties of A. kunkeei EABW06, isolated from beeswax, to assess its potential for human and biotechnological applications. A. kunkeei EABW06 demonstrated gastrointestinal resilience, with > 96% survival at pH 3.0 and tolerance to 0.3-0.7% bile salts. It exhibited broad-spectrum antimicrobial activity against Escherichia coli O157:H7, Salmonella typhi, and Clostridioides difficile. Additionally, A. kunkeei EABW06 inhibited fungal pathogens including Aspergillus fumigatus, Aspergillus flavus, and Candida albicans. HPLC analysis revealed acetic, lactic, butyric, and propionic acids as dominant metabolites, correlating with its antimicrobial effects. Safety assessments confirmed no hemolysis, non-cytotoxicity to Caco-2 cells, and sensitivity to clinically relevant antibiotics. In milk fermentation trials, A. kunkeei EABW06 reduced pH to 4.5 ± 0.1, produced 0.98 ± 0.03% lactic acid (titratable acidity), and exhibited proteolytic activity (0.384 ± 0.002) measured by spectrophotometric quantification. In conclusion, A. kunkeei EABW06 is a safe, robust probiotic candidate with potential applications in biopreservation and dairy biotechnology.