Assessment and Optimization of Methods for the Detection of <i>Flavobacterium</i> Phages at Low Concentrations in Water Samples

Background: The development of phage libraries is vital for phage-based applications and to further the understanding of phage–host interactions. However, phage isolation from environmental samples on a large scale is resource-intensive. Methods: To optimize the detection of phages from environmental water samples with low phage concentrations, we assessed and optimized a small-volume enrichment method using the important fish pathogen Flavobacterium psychrophilum as a target host. Results: The 96-well plate enrichment assay demonstrated phage activity in samples with initial phage concentrations of a single plaque-forming unit (PFU) per well, and the overall detection limit of the method was 0.011 PFU mL −1 in the water sample. For phage concentrations &gt;10 PFU, the phage concentration in the sample could be predicted from optical density inhibition during incubation. When applied to 45 water samples collected from a recirculating aquaculture system, phages were detected in 36 of the samples. Conclusion: While this method provides resource-efficient means to isolate phages from aquatic environments, phage–host system-dependent variations are shown, and optimization for other phage–host systems is recommended.