Introduction
Diseases caused by pathogenic vibrios cause major losses in aquaculture. The use of antibiotics in order to control these infections has led to the development and spread of antibiotic-resistant pathogens, rendering antibiotic treatments ineffective and causing problems for food safety (Defoirdt et al., 2011). Therefore, novel methods to control vibriosis are needed. A promising alternative is the interference with quorum sensing, bacterial cell-to-cell communication (Defoirdt, 2018). Recent research has shown that indole controls virulence-related phenotypes in pathogenic bacteria and i ndole is one of the cell-to-cell signaling molecules produced by vibrios. Consequently, we aimed at investigating the interference with indole signaling in order to control vibriosis in aquaculture.
Results and Discussion
We found that indole controls several virulence-related phenotypes in vibrios, most notably biofilm formation and motility . Further , we found that the addition of 100-200 µM indole decreases the virulence to aquatic organisms in all marine vibrios studied thus far, including V. anguillarum (in sea bass), V. campbellii (in brine shrimp and giant river prawn), V. crassostreae (in blue mussel), V. parahaemolyticus (in brine shrimp) and V. tasmaniensis (in blue mussel) (Li et al., 2014; Yang et al., 2017; Zhang et al., 2022c; Zhang e t al., 2022d). At these concentrations, indole did not affect the growth of the vibrios. This is important because it indicates that there will be a lower risk for the spread of resistance against the virulence-decreasing effect of indole signaling when compared to antibiotics.
Given the fact that indole signaling controls the virulence of Vibrio species, we have been searching for more potent indole analogues to control vibriosis. I ndole analogues are widely present in nature (Lee et al., 2015). Most notably are the auxin plant hormone indole-3-acetic acid and its precursors such as indole-3-acetamide and indole-3-acetonitrile (Zhao, 2010). Indole-3-acetic acid has been shown to decrease biofilm formation and motility of V. campbellii , V. harveyi and V. parahaemolyticus strains at 200 µM and to protect brine shrimp larvae from these pathogens when added at 400 µM to the rearing water (Zhang et al., 2023). The auxin precursors indole-3-acetamide and indole-3-acetonitrile were shown to have a similar effect as indole-3-acetic acid, with indole-3-acetonitrile being active at a relatively low concentration of 10 µM (Yang et al., 2017; Zhang et al., 2022b). Interestingly, auxins are not only produced by terrestrial plants, but also by algae and seaweeds (Stirk et al., 2004; Lin et al., 2022). Hence, micro-algae and seaweeds could be interesting sources of auxins to control vibriosis in aquaculture.
In addition to natural indole analogues, we have also studied synthetic derivatives. A first group of derivatives are indene, 2,3-benzofuran and thianaphthene, in which the N atom of indole is replaced by C, O and S, respectively. All three of these compounds were found to increase the survival of brine shrimp larvae challenged with V. campbellii to around 80% or more when added to the rearing water at 200 µM. Further, they significantly decreased swimming motility, but had no effect on biofilm formation. A second interesting group of indole analogues are halogenated indoles. We investigated the impact of 31 halogenated indoles on V. campbellii (Zhang et al., 2022a) . None of the compounds affected growth of V. campbellii for concentrations up to 200 µM, whereas 10 compounds increased the survival of brine shrimp challenged with V. campbellii to over 80% when added to the rearing water at 20 µM, and 5 compounds (6-bromoindole, 7-bromoindole, 4-fluoroindole, 5-iodoindole and 7-iodoindole) did so when added at 10 µM. All of these compounds decreased swimming motility of V. campbellii at 10 µM and most of them inhibited biofilm formation at 100 µM.
In conclusion, we found that indole signaling controls the virulence of all vibrios we have tested thus far in different host organisms (fish, bivalves and crustaceans) and we identified several indole analogues that are highly effective in blocking the virulence of vibrios. These results indicate that indole signaling is an interesting target for the control of vibriosis in aquaculture.
References
Defoirdt, T., 2018. Quorum-sensing systems as targets for antivirulence therapy. Trends Microbiol . 26, 313-328.
Defoirdt, T., Sorgeloos, P., and Bossier, P., 2011. Alternatives to antibiotics for the control of bacterial disease in aquaculture. Curr. Opin. Microbiol. 14, 1-8.
Lee, J.H., Wood, T.K., Lee, J., 2015. Roles of indole as an interspecies and interkingdom signaling molecule. Trends Microbiol. 23, 707-718.
Li, X., Yang, Q., Dierckens, K., Milton, D.L., Defoirdt, T., 2014. RpoS and indole signaling control the virulence of Vibrio anguillarum towards gnotobiotic sea bass (Dicentrarchus labrax ) larvae. PLoS One 9, e111801.
Lin, H., Li, Y., Hill, R.T., 2022. Microalgal and bacterial auxin biosynthesis: implications for algal biotechnology. Curr. Opin. Biotechnol. 73, 300-307.
Stirk, W.A., Arthur, G.D., Lourens, A.F., Novak, O., Strnad, M., van Staden, J., 2004. Changes in cytokinin and auxin concentrations in seaweed concentrates when stored at an elevated temperature. J. Appl. Phycol. 16, 31-39.
Yang, Q., Pande, G.S.J., Wang, Z., Lin, B., Rubin, R.A., Vora, G.J., Defoirdt, T., 2017. Indole signalling and (micro)algal auxins decrease the virulence of Vibrio campbellii , a major pathogen of aquatic organisms. Environ. Microbiol . 19, 1987-2004.
Zhang, S., Yang, Q., Defoirdt, T., 2022a. Halogenated indoles decrease the virulence of Vibrio campbellii in a gnotobiotic brine shrimp model. Microbiol. Spectrum 10, e02689-22.
Zhang, S., Yang, Q., Defoirdt, T., 2022b. Indole analogues decreasing the virulence of Vibrio campbellii towards brine shrimp larvae. Microbial Biotechnol. 15, 2917-2928.
Zhang, S., Yang, Q., Defoirdt, T., 2022c. Indole decreases the virulence of pathogenic vibrios belonging to the Harveyi clade. J. Appl. Microbiol. 132, 167-176.
Zhang, S., Yang, Q., Fu, S., Janssen, C.R., Eggermont, M., Defoirdt, T., 2022d. Indole decreases the virulence of the bivalve model pathogens Vibrio tasmaniensis LGP32 and Vibrio crassostreae J2_9. Sci. Rep. 12, 5749.
Zhang, S., Van Haesebroeck, J., Yang, Q., Defoirdt, T., 2023. Indole-3-acetic acid increases the survival of brine shrimp challenged with vibrios belonging to the Harveyi clade. J. Fish Dis . 46, 477-486 .
Zhao, Y.D., 2010. Auxin biosynthesis and its role in plant development. Annu. Rev. Plant Biol. 61, 49-64.