Bioactive compounds in fluid propolis preparations inhibit different life stages of pathogenic oomycetes Aphanomyces astaci and Saprolegnia parasitica

Abstract

The pathogenic oomycetes Saprolegnia parasitica, causative agent of saprolegniosis in salmonid fish, and Aphanomyces astaci, causative agent of crayfish plague, have a negative impact on freshwater aquaculture. As they are controlled worldwide with chemicals that are harmful to humans and the environment, there is a growing need to replace them with ecologically acceptable alternatives. Propolis has documented antimicrobial activity as well as positive effects on the immune response, growth performance and/or reproductive capacity of fish and crayfish. Therefore, we aimed to investigate the possible inhibition of S. parasitica and A. astaci by fluid propolis formulations. Chemical analysis showed that the propolis formulations were rich in volatile and non-volatile phenolic bioactive components (chrysin was the dominant flavone and pinocembrin the dominant flavanone). Overall, the fluid propolis formulations showed good in vitro inhibition of mycelial growth and zoospores of both pathogens, but we found differences in sensitivity depending on species and life cycle stage. Mycelial growth was more sensitive for A. astaci, with EC50 values (the samples concentrations causing 50% inhibition) up to 8.59 μg/mL, compared to 206.60 μg/mL for S. parasitica. Zoospore motility was more affected in S. parasitica, where MIC (minimum inhibitory concentration) values were up to 61.88 μg/mL, compared to 154.68 μg/mL for A. astaci. Zoospore germination of both pathogens was similarly sensitive to the fluid propolis preparations, with EC50 values of up to 19.52 μg/mL for A. astaci and up to 23.62 μg/mL for S. parasitica. In addition, molecular docking was used to analyze the binding of selected propolis components to oomycete proteins suggested to play a role in pathogenesis. Apigenin, chrysin and pinocembrin were predicted to bind strongly to the endochitinase of A. astaci, which is mainly expressed in the mycelium, and to the thrombospondin of S. parasitica, mainly expressed in the cysts, which is consistent with the results of the in vitro inhibition experiments. Overall, our results suggest that propolis could be used in salmonid and crayfish aquaculture not only as an immunostimulant but also as an antioomycetic agent.