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.
