Isolation, in vitro characterization and efficacy assessment in Galleria mellonella larvae of four bacteriophages targeting Aeromonas salmonicida

The Gram-negative bacteria Aeromonas (A.) salmonicida is a primary fish pathogen that causes furunculosis in salmonids as well as septicemia in a variety of fish. In one hand because this disease is responsible for significant losses in salmonid production worldwide and in the other hand because of the frightening tendency of this bacteria to exhibit antimicrobial (multi-)resistances, phage therapy could represent a leading alternative to treat this infection in aquaculture. The aims of this study were to create a collection of A. salmonicida strains, isolate phages targeting these strains, phenotypically and genomically characterize these newly isolated phages and finally assess their potential for phage therapy in the preliminary in vivo model of Galleria (G.) mellonella larvae. Four new phages active against A. salmonicida were isolated from water samples collected in fish farms and natural aquatic environments in southern Belgium. Genomic analysis showed that 3 of these phages, named vB_AsaM_ULASA2 (170,823bp), vB_AsaM_ULASA3 (164,381bp) and vB_AsaM_ULASA4 (171,205bp), belong to the Straboviridae family while vB_AsaM_ULASA1 (47,813bp) stay in the unclassified part of the Caudoviricetes class. All 4 presented a myovirus morphotype. Four-day efficacy experiments in the preliminary in vivo model of G. mellonella larvae showed that 3 of these 4 phages were responsible for a significant extension in the larval survival time at the 2 treatment doses tested (MOI 10 and 100). In light of these results, these phages targeting A. salmonicida could represent potential new candidates for the development of anti-furunculosis phage treatments in aquaculture.