Bacteriophage isolation using enrichment cultures
Potential host cells are grown overnight in liquid cultures containing a rich growth medium ( e . g . , MLB ) .
Adjust to an optical density measured at 525 nm ( OD525 ) of 0.3–0.5 .
Approximately 25 mL water sample is filtered ( 0.2 µm or 0.45 µm syringe filters ) to minimize the risk of bacterial contamination of the enrichment cultures .
Transfer filtered samples to triplicate sterile 100 mL culture flasks .
Add 3 mL 10× growth medium ( i . e . , 10 times concentrated medium ) .
Start enrichment cultures by adding 1 mL of each of the target host strains of interest ( e . g . , 1–6 different strains for each incubation ) to the culture flask .
Incubate the cultures on a shaking table at a temperature and period that is appropriate for the host bacteria .
A control culture is established where the environmental sample is replaced by 25 mL artificial seawater ( or 0.02 µm filtered water sample ) to verify bacterial growth in absence of phages .
