ENVIRONMENTAL BIOTECHNOLOGY Quorum sensing-disrupting coumarin suppressing virulence phenotypes in Vibrio splendidus Shanshan Zhang 1 & Ningning Liu 1 & Weikang Liang 1 & Qingxi Han 1 & Weiwei Zhang 1 & Chenghua Li 1 Received: 12 July 2016 /Revised: 6 November 2016 /Accepted: 9 November 2016 # Springer-Verlag Berlin Heidelberg 2016 Abstract In the present study, the effects of an environmental friendly natural reagent coumarin, on the growth and potential virulence factors, as well as its ability to interfere the infection of Vibrio splendidus (Vs), were determined. Coumarin showed no effects on the maximal growth of Vs, and biofilm formation of Vs, while it significantly decreased protease ac- tivity and hemolytic activity by 43 and 80%, respectively. Correspondingly, coumarin exhibited an obviously protective effect, with a relative percent survival of 60% upon Apostichopus japonicus from infection by Vs. To preliminar- ily investigate the mechanism underlining the inhibitory ef- fects, regulation of genes Vsm and Vsh respectively related to protease activity and hemolytic activity by supernatant and supernatant extract containing acyl-homoserine lactones (AHLs) and coumarin was determined. Cell-free supernatant from higher density and its ethyl acetate extract containing AHL signal molecules could respectively upregulate the mRNA level of Vsm by 17.4- and 2.3-fold and Vsh by 7.2- and 5.0-fold, when Vs was at lower cell density. However, coumarin could reduce the stimulatory effects of both the su- pernatant and its ethyl acetate extract. Combining all the results in our study, it was suggested that coumarin could be consid- ered as an alternative to be used for controlling infection of Vs, downregulating the expression of potential virulence factors through interfering the AHL-mediated pathways. Keywords Coumarin . Vibrio splendidus Vs . Protease . Hemolytic activity . Protective effect Introduction The invertebrate sea cucumber Apostichopus japonicus (Echinodermata, Holothuroidea) is an economically important species in Chinese marine culture (Liu et al. 2010a); however, with its rapid expansive and intensive farming, skin ulceration syndrome (SUS), manifesting swollen mouth, viscera ejec- tion, skin ulceration, and massive mortality have resulted from pathogenic infection, leading to huge economical loss (Deng et al. 2008; Li et al. 2010). Vibrio splendidus is one of the most important opportunistic pathogens that could infect A. japonicus, leading to the outbreak of SUS (Zhang et al. 2006). Thus, ecological strategies with the merits of high ef- ficiency and environmental friendly to protect A. japonicus from infection by V. splendidus have become urgently recom- mended and have become prior choices, compared with the traditional methods of using antibiotics and chemotherapeu- tics, which own the shortage of drug-resistant development, environmental pollution, and residues in the environment (Zhang et al. 2010; Li et al. 2015). Coumarin is derived from plants such as Rutaceae, Umbelliferae, Asteraceae, Leguminosae, Thymelaeaceae, and Solanaceae, and its pharmacological safety has been established. Small dosage of coumarin (≤0.64 mg/kg BW/ day) in food or cosmetics does not pose any risk to human health (Felter et al. 2006). Thus, coumarin has been suggested as a powerful alternative for disease control, as it is determined to be an antibacterial reagent and a universal quorum sensing inhibitor (QSI) (Gutiérrez-Barranquero et al. 2015). The anti- microbial activities of coumarin had been detected in the leaf extracts of Petroselinum crispum and Ruta graveolens (Ojala * Weiwei Zhang zhangweiwei1@nbu.edu.cn * Chenghua Li lichenghua@nbu.edu.cn 1 School of Marine Sciences, Ningbo University, 818 Fenghua Road, Ningbo, Zhejiang Province 315211, People’ s Republic of China Appl Microbiol Biotechnol DOI 10.1007/s00253-016-8009-3