In screening the culture broth of marine bacteria collected at Yap (Micronesia), Palau (Belau), and Okinawa (the southwest islands of Japan) for antimicroalgal activity, 37 out of 2,594 bacterial isolates tested were found to produce anticyanobacterial substances against Oscillatoria amphibia NIES-361. One strain, C-979, identified as a Vibrio sp., was selected and cultured in 2.4 liters of marine broth 2216 to identify the bioactive compound produced by the strain. The purified very hydrophilic compound (16.4 mg) was determined to be -cyano-L-alanine (LCNAla) by instrumental analyses and the application of the advanced Marfey method. L-CNAla did not inhibit the growth of bacteria, yeast, or eukaryotic microalgae, but some cyanobacteria were found to be sensitive to L-CNAla at a concentration of 0.4 to 25 g/ml. The effect of L-CNAla on some other environmental organisms, including invertebrates and a macroalgae, is discussed. CNAla production in marine broth was examined by thin-layer chromatography for the 37 bacterial isolates which produced an anticyanobacterial substance. The broth of 36 of these strains contained CNAla, suggesting the wide distribution of CNAla production by marine bacteria. This is the first report on bacteria that produce CNAla without a supply of the cyanide ion in the medium.Research studies of inhibitors of bacteria, fungi, and cultured cells have accumulated, and many effective drugs have appeared for clinical use. However, the number of reports of antimicroalgal compounds is relatively small, although these organisms cause such problems as blooms by cyanobacteria, red tide and the production of marine toxins by dinoflagellates, and biofilm formation on marine structures by diatoms. In particular, some bloom-forming cyanobacteria are known to produce toxic metabolites such as anatoxins (8) and microcystins (3). The mechanism by which the cyanobacteria form blooms remains to be investigated, but the threat and damage to human and animal life and to industry are serious. In the present study, we screened strains for antimicroalgal compounds against one cyanobacterium and three eukaryotic microalgae. A culture broth of each of 2,594 marine bacterial strains was examined in this screening, and among them, 37 strains were found to produce anticyanobacterial substances in the culture on marine broth 2216 (MB). Interestingly, no inhibitory activity toward the three eukaryotic microalgae tested was apparent in the broths of any of these bacterial strains.We purified the anticyanobacterial compound from C-979, which showed the highest anticyanobacterial activity. The chemical structure was determined for the compound, and the bioactivity profile was examined. Furthermore, we examined if the above-mentioned 37 strains produce the same anticyanobacterial compound that C-979 produces.
MATERIALS AND METHODSMicroalgae and cultivation medium. All the microalgal strains and the media used in this study are listed in Table 1. F/2, K ϩ ESM, MC, SOT, and CT media have been described elsewhere by Guilla...