Vibriophages Differentially Influence Biofilm Formation by Vibrio anguillarum Strains

Abstract

Vibrio anguillarum is an important pathogen in marine aquaculture, responsible for vibriosis. Bacteriophages can potentially be used to control bacterial pathogens; however, successful application of phages requires a detailed understanding of phage-host interactions under both free-living and surface-associated growth conditions. In this study, we explored in vitro phage-host interactions in two different strains of V. anguillarum (BA35 and PF430-3) during growth in microcolonies, biofilms, and freeliving cells. Two vibriophages, ⌽H20 (Siphoviridae) and KVP40 (Myoviridae), had completely different effects on the biofilm development. Addition of phage ⌽H20 to strain BA35 showed efficient control of biofilm formation and density of free-living cells. The interactions between BA35 and ⌽H20 were thus characterized by a strong phage control of the phage-sensitive population and subsequent selection for phage-resistant mutants. Addition of phage KVP40 to strain PF430-3 resulted in increased biofilm development, especially during the early stage. Subsequent experiments in liquid cultures showed that addition of phage KVP40 stimulated the aggregation of host cells, which protected the cells against phage infection. By the formation of biofilms, strain PF430-3 created spatial refuges that protected the host from phage infection and allowed coexistence between phage-sensitive cells and lytic phage KVP40. Together, the results demonstrate highly variable phage protection mechanisms in two closely related V. anguillarum strains, thus emphasizing the challenges of using phages to control vibriosis in aquaculture and adding to the complex roles of phages as drivers of prokaryotic diversity and population dynamics. Vibrio anguillarum is a marine pathogenic bacterium causing vibriosis, a fatal hemorrhagic septicemia, which contributes to significant mortalities in fish and shellfish aquaculture worldwide (1-3). The persistence of Vibrio pathogens in aquaculture has been attributed to their ability to form biofilms with increased tolerance of disinfectants and antibiotics (4, 5). Moreover, the first stage of infection involves biofilm-like microcolonies in the skin tissue, causing chronic infection (5).Recently, bacteriophages have been suggested as potential agents of pathogen control in aquaculture, and the controlling effects of phages have been explored for a number of fish pathogens (6-8). Successful application of phages to reduce vibriosis-related mortality has been demonstrated (9, 10). The capabilities of some phages to produce depolymerases, which hydrolyze extracellular polymers in bacterial biofilms, have made the use of bacteriophages particularly relevant in the treatment of biofilm-forming pathogens, as demonstrated in biofilms of Pseudomonas aeruginosa (11), Escherichia coli (12), and Staphylococcus aureus (13).Apart from the potential physical and chemical barrier provided by biofilms, the use of bacteriophages to control pathogens is challenged by the development of and selection for phage-resistant or phage-t...

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Isolation and Characterization of a Shewanella Phage–Host System from the Gut of the Tunicate, Ciona intestinalis

“…Previously, Vibrio anguillarum phages have been shown to have different influences on biofilm formation over long-term cultures, due mostly to varied microcolony morphologies with one strain having flat single layers (BA35), and the other forming complex 3D structures (PF430-3) [87]. This structural distinction resulted in differential penetrability of the biofilm structure by phages during the initial stages of biofilm formation, and ultimately impeded the formation of the BA35 single layer biofilm compared to the control.…”

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