Exopolysaccharide-producing isolates from Thai milk kefir and their antioxidant activities

• Only Bacillus spp. were found in Thai milk kefir with predominant Bacillus amyloliquefaciens.
• All Bacillus spp. were able to produce exopolysaccharide (EPS) on glucose, lactose and sucrose.
• Selected strains with EPS exhibited resistance to H2O2 and antioxidant activity.
• Cells with de-structured or removed EPS showed reduced bioactivity.

This is the first report of microbial diversity in milk kefir from Thailand and isolates with exopolysaccharide (EPS)-producing capacity. Microbes in milk kefir were isolated on selective agars and the purified bacteria were identified using PCR-based 16S rDNA gene analysis. Twenty four strains of Bacillus spp. were identified out of 85 isolates without any lactic acid bacteria, acetic acid bacteria or yeasts. Closest relative isolates of Bacillus amyloliquefaciens SD-32 (100% identity) were predominant (67% abundance) followed by Bacillus sp. DH25 (95% identity, 7% abundance), uncultured Bacillus sp. clone L4 (95% identity, 5% abundance) and the other 21 Bacillus spp. each accounts for only 1% abundance. All Bacillus spp. identified were able to produce EPS on MRS agar with 8% sugar (sucrose, lactose or glucose) using the disk diffusion method suggesting their capacity to utilise sucrose, lactose and glucose as precursors to EPS production however EPS appearances varied depending upon bacterial strain and type of sugar. Selected bacteria grown in 8% sucrose exhibited 18–27% DPPH scavenging activity and resistance to H2O2. Removal of EPS showed significantly reduced bioactivity DPPH scavenging activity and resistance to H2O2. Microbial diversity of Thai milk kefir was unique from other milk kefirs of different origins.