EVALUATE THE EFFECT OF PHAGES USE IN ENVIRONMENTAL BACTERIAL ECOLOGY EVALUAR EL USO DE FAGOS EN LA ECOLOGÍA BACTERIANA AMBIENTAL

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1 Coordinator Partners universidade de aveiro ENVIPHAGE. LIFE13 ENV/ES/ EVALUATE THE EFFECT OF PHAGES USE IN ENVIRONMENTAL BACTERIAL ECOLOGY EVALUAR EL USO DE FAGOS EN LA ECOLOGÍA BACTERIANA AMBIENTAL AVALIAR O EFEITO DE FAGOS NA ECOLOGIA BACTERIANA AMBIENTAL LIFE13 ENV/ES/ LAYMAN S R E P O R T

2 Background Aquaculture is a growing sector that generates significant economic activity and provides society with high-demand food products. However, fish farming is facing problems due to the concentration of fish and organic matter in limited spaces, which sometimes causes disease transmission in facilities. Although different strategies have been developed to fight bacterial infections, few of them are applicable in spawning stages and with small juvenile fish. For the most part, these treatments include antibiotics, but there are problems with their use: it is controlled; consumers request antibiotic-free products; and they may cause bacteria to become resistant, which could become a public health problem. Bacteriophages (or phages) are organisms that infect and destroy specific bacteria. In recent years, bacteriophages have been suggested as an alternative to antibiotics in aquaculture. Very promising results have been obtained in laboratories, but for their use at an industrial level we need to know about the environmental impact of phages, especially in bacterial ecology, before proposing their regular use in the aquaculture industry. This project seeks to address this gap between the laboratory and industrial-scale treatment.

3 Goals The LIFE-Enviphage project aims to assess the environmental effects of the controlled use of bacteriophages in an aquaculture production system. Specifically, the goals are: To evaluate the effect of bacteriophages on the microorganisms naturally present in the waters of fish farms and in the surrounding open waters. To evaluate the effect of bacteriophages on the bacteria of the fish bred in fish farms. To study the effect of bacteriophages on animal models as a way of predicting their behaviour in humans.

4 Main activities Based on genetic technologies, this project will study the effect of bacteriophages in environmental and intestinal bacteria communities. Evaluation of the impact of the use of bacteriophages on bacterial populations present in the fish farm waters and in surrounding waters. Isolation, selection, characterisation and production of bacteriophages against pathogenic bacteria in aquaculture. Assessment of the effect of the use of bacteriophages on the intestinal microbiota of commercial fish. Study of the effect of phages using an animal model. Study of their immunostimulating capacity on an animal model and modelling of their effect on human intestinal bacteria. Economic assessment of phage technology for pathogen control in aquaculture.

5 Main results obtained 1. No risks have been identified in their use in animal models. Bacteriophages do not stimulate the genes associated with the immune response in fish models, which suggests that the immune system of the animal does not recognise the phages as possible pathogens, avoiding the consequences of an immune response. 2. Bacteriophages do not significantly affect bacterial ecosystems. The evolution of microorganisms has been studied in four environments that are susceptible to bacteriophage exposure: fish farm waters, waters surrounding fish farming facilities, the intestinal tract of exposed fish and a bacterial model of human flora in animal models. No drastic effect of the bacteriophages on the bacterial ecosystems was observed, and no variations beyond the natural variations of the ecosystems were observed. 3. Bacteriophages are a viable alternative. During the project, bacteriophages to fight pathogens in aquaculture were isolated, selected, studied, produced and tested successfully in real conditions. This project demonstrates that real-scale phage production is a fast, effective, safe and viable alternative to antibiotics, reducing the time needed to put a specific antibacterial product on the market.

6 Transferable results Environmental assessment protocol Over the course of the project, a systematic protocol was introduced to ascertain the environmental impact of the bacteriophages produced. The experiment presented in the project can be taken as an example for future developments. Local authorities have a series of recommendations to assess the effect of phages on the environment and on the microorganisms of the human intestinal tract.

7 Results The results show that the use of AS-P1 bacteriophages does not have any implications for the environment, this is an effective alternative for reducing/eliminating the load of Aeromonas salmonicida in aquaculture plants and their production is possible at the industrial level. Furthermore, the models used suggest that this bacteriophage does not have any effects on human health. Lastly, the results show that industrial-scale production of bacteriophages is currently a viable technology. Collections Over the course of the project, two bacteriophage collections active against pathogens in aquaculture have been created. With these collections, fish farmers have a tool available to eliminate fish pathogens without any consequences for the environment, fish, microorganisms or humans. This would enormously reduce their environmental impact, while increasing the profitability of farms by lowering mortality in early stages of the breeding process.

8 EVALUATE THE EFFECT OF PHAGES USE IN ENVIRONMENTAL BACTERIAL ECOLOGY EVALUAR EL USO DE FAGOS EN LA ECOLOGÍA BACTERIANA AMBIENTAL AVALIAR O EFEITO DE FAGOS NA ECOLOGIA BACTERIANA AMBIENTAL LIFE13 ENV/ES/001048