AN IMPROVED PROTOCOL FOR TESTING ESSENTIAL OIL ACTIVITY ON FISH PATHOGEN BIOFILM

Aquaculture is recognized as the fastest-growing food-producing industry globally for human consumption (Naylor et al., 2022). The European aquaculture sector alone yields more than 3 million tons of fish yearly, as reported by the Food and Agriculture Organization (FAO, 2020). Numerous marine and freshwater aquatic organisms are vulnerable to bacterial diseases such as vibriosis and photobacteriosis, yersiniosis and aeromonosis respectively, just to mention a few very common ones. The origin of fish pathogens can be correlated to various factors, but their ability to withstand and cause disease outbreaks in aquaculture systems suggests that biofilms are a possible source of persistence, especially for multidrug-resistant bacterial strains that have adapted to the facility settings and may serve as the main source of infection for farmed fish. Therefore, the capacity to develop a biofilm represents a significant concern with possible implications in disease outbreaks as a pathogen reservoir as already demonstrated for some bacterial agents of importance in aquaculture such as Yersinia ruckeri (Coquet et al., 2002). Microbial cells can adhere to each other and to host cells and abiotic surfaces, such as glass, polystyrene plastic, and seashells which promotes colonization and the formation of biofilms. The biofilm comprises densely populated bacteria shielded by a robust exopolymer matrix that firmly attaches to a surface. Forming biofilms leads to the failure of antimicrobial agents, with 1000-fold greater resistance to them (Uruen et al., 2020). The main objective in counteracting fish pathogen infections is the identification of substances (possibly of natural origin, e.g., essential oils), which can effectively target pathogen biofilms. The first step of this approach is the study of in vitro systems, which help to demonstrate the efficacy of the selected antibiofilm agents. The present work aimed to improve the currently available techniques to obtain a formed biofilm by fish pathogens as the first step in studying antibiofilm substances.