Phage-Mediated Control of Flavobacterium psychrophilum in Aquaculture: In vivo Experiments to Compare Delivery Methods

Phage-based approaches have gained increasing interest as sustainable alternative strategies to antibiotic treatment or as prophylactic measures against disease outbreaks in aquaculture. The potential of three methods (oral, bath, and injection) for delivering a two-component phage mixture to rainbow trout fry for controlling Flavobacterium psychrophilum infections and reduce fish mortality was investigated using bacteriophages FpV4 and FPSV-D22. For the oral administration experiment, bacteriophages were applied on feed pellets by spraying (1.6 × 10⁸ PFU g^–1) or by irreversible immobilization (8.3 × 10⁷ PFU g^–1), using the corona discharge technology (Fixed Phage Ltd.). The fish showed normal growth for every group and no mortality was observed prior to infection as well as in control groups during the infection. Constant detection of phages in the intestine (∼10³ PFU mg^–1) and more sporadic occurrence in kidney, spleen, and brain was observed. When fish were exposed to F. psychrophilum, no significant effect on fish survival, nor a direct impact on the number of phages in the sampled organs, were detected. Similarly, no significant increase in fish survival was detected when phages were delivered by bath (1^st and 2^nd bath: ∼10⁶ PFU ml^–1; 3^rd bath: ∼10⁵ PFU ml^–1). However, when phages FpV4 and FPSV-D22 (1.7 × 10⁸ PFU fish^–1) were administered by intraperitoneal injection 3 days after the bacterial challenge, the final percent survival observed in the group injected with bacteriophages FpV4 and FPSV-D22 (80.0%) was significantly higher than in the control group (56.7%). The work demonstrates the delivery of phages to fish organs by oral administration, but also suggests that higher phage dosages than the tested ones may be needed on feed pellets to offer fish an adequate protection against F. psychrophilum infections.