Effects of antifungal agents on red tide phytoplankton: possibility of a novel preventive tool

Abstract

Harmful algal blooms (HABs) are having signi¢cant impacts on coastal and estuarine areas, aiecting the health of aquatic organisms and population growth, ecology and even humans (Anderson1997). Because the HABs often cause acute shell¢sh poisonings and mass mortalities of aquatic organisms depending on the causative species of HABs, the impacts of HABs on aquatic industries and economies are serious problems (Landsberg 2002). Aquaculture is an important industry as a source of seafood. Aquaculture operations are carried out around coastal areas, and also red tides occurring near the shore frequently cause mass mortality of cultured ¢sh. Chattonella marina (Raphidophyceae) and Heterocapsa circularisquama (Dino£agellata) are known to have lethal eiects on ¢sh and shell¢sh, respectively (Matsuyama 1999; Landsberg 2002). There is no eiective prevention against causative red tide phytoplankton. In our previous study, a useful marine bacterium was isolated during screening for antifungal compounds (Nakashima, Kurachi, Kato,Yamaguchi & Oda 2005; Nakashima, Kato,Yamaguchi & Oda 2005). This marine bacterium produced a prodigiosin analogue that had antifungal activity against dermatophytes (Nakashima, Kato et al. 2005). In addition, the prodigiosin analogue also showed potent algicidal activity against red tide phytoplankton (Nakashima, Miyazaki, Matsuyama, Muraoka, Yamaguchi & Oda 2006). The phytoplankton may have common target sites that antifungal agents act on dermatophytes. It is possible that antifungal agents are novel preventive materials against red tide phytoplankton. Therefore, to con¢rm the hypothesis, we investigated the eiects of commercial antifungal agents (Fig. 1) on red tide phytoplankton. Bifonazole (BFZ), terbina¢ne hydrochloride (TBF), cholesterol and ergosterol (Fig. 1) were purchased fromWako Pure Chemical Industries (Tokyo, Japan). Stock solutions of antifungal agents were prepared with dimethyl sulfoxide to a concentration of 10mgmL . Axenic cultures of C. marina and H. circularisquamawere maintained at 26 1C in Erd-Schreiber modi¢ed (ESM) medium (pH 8.2) under illumination from a £uorescent lamp (30 mmol photonsm 2 S ) with a cycle of 12 h light and 12 h dark as described previously (Nakashima et al. 2006). To determine the algicidal activities of antifungal agents against C. marina and H. circularisquama, the stock solution of each antifungal agent (10mgmL ) was diluted 500-fold in ESMmedium, and serial twofold dilutions in ESM medium were prepared. Red tide phytoplankton in the logarithmic growth phase were diluted with ESM medium to a density of 2 10 cells mL . ESM media (500 mL) with various concentrations of antifungal agent were added to each algal cell suspension (500 mL) in the wells of a 24-well plate.The ¢nal concentrations of antifungal agents ranged from 10 to 0.01 mgmL . After incubating at 26 1C for 24 h, the number of viable algal Aquaculture Research, 2008, 39, 1346^1350 doi:10.1111/j.1365-2109.2008.01998.x