Reinforced clostridial broth

Bacterial strains representing species commonly found in bovine milk were used to construct a mock bacterial community (Table 1). Each bacterial strain was grown in standard laboratory culture medium with negative controls for that species and harvested at early stationary phase by centrifugation at 13,000 × g for 2 min. The laboratory culture media were as follows: Bacillus subtilis, Pseudomonas fluorescens, and Escherichia coli, LB (Lennox broth; Thermo Fisher Scientific); Enterococcus faecalis and Streptococcus agalactiae, brain heart infusion broth (Thermo Fisher Scientific); Staphylococcus aureus, tryptic soy broth (Becton Dickinson); Corynebacterium bovis, tryptic soy broth (Becton Dickinson) with 0.1% Tween 80; Lactococcus lactis, M17 broth (Becton Dickinson) with 0.5% glucose; and Clostridium tyrobutyricum, reinforced clostridial broth (Becton Dickinson). All strains were incubated at 37°C, with the exception of B. subtilis, L. lactis, and P. fluorescens, which were incubated at 30°C. B. subtilis, C. bovis, E. faecalis, E. coli, and P. fluorescens were grown under aeration (250 rpm).

Bacterial strains from species commonly found in milk and other dairy products were selected for a BCMC (Table 1). The strains were grown as previously described [9 (link)] in the following culture media: LB (Lennox broth; Thermo Fisher Scientific, Waltham, WA, USA) for Bacillus subtilis S44, Pseudomonas fluorescens A506, and Escherichia coli ATCC 700728; brain heart infusion broth (Thermo Fisher Scientific, Waltham, WA, USA) for Enterococcus faecalis ATCC 29212 and Streptococcus agalactiae ATCC 27956; tryptic soy broth (Becton Dickinson, Franklin Lakes, NJ, USA) for Staphylococcus aureus ATCC 29740 and Corynebacterium bovis ATCC 7715; M17 broth (Becton Dickinson, Franklin Lakes, NJ, USA) with 0.5% w/v glucose for Lactococcus lactis IL1403; and reinforced clostridial broth (Becton Dickinson, Franklin Lakes, NJ, USA) for Clostridium tyrobutyricum ATCC 25755.