An 80% methanolic extract of Rhus verniciflua Stokes bark showed significant anti-viral activity against fish pathogenic infectious hematopoietic necrosis virus (IHNV) and viral hemorrhagic septicemia virus (VHSV) in a cell-based assay measuring virus-induced cytopathic effect (CPE). Activity-guided fractionation and isolation for the 80% methanolic extract of R. verniciflua yielded the most active ethyl acetate fraction, and methyl gallate (1) and four flavonoids: fustin (2), fisetin (3), butin (4) and sulfuretin (5). Among them, fisetin (3) exhibited high antiviral activities against both IHNV and VHSV showing EC₅₀ values of 27.1 and 33.3 μM with selective indices (SI = CC₅₀/EC₅₀) more than 15, respectively. Fustin (2) and sulfuretin (5) displayed significant antiviral activities showing EC50 values of 91.2–197.3 μM against IHNV and VHSV. In addition, the antiviral activity of fisetin against IHNV and VHSV occurred up to 5 hr post-infection and was not associated with direct virucidal effects in a timed addition study using a plaque reduction assay. These results suggested that the bark of R. verniciflua and isolated flavonoids have significant anti-viral activity against IHNV and VHSV, and also have potential to be used as anti-viral therapeutics against fish viral diseases.

Our previous investigation revealed that 80% methanolic extract of the red alga Polysiphonia morrowii has significant antiviral activities against fish pathogenic viruses, infectious hematopoietic necrosis virus (IHNV) and infectious pancreatic necrosis virus (IPNV). The present study was conducted to identify compounds attributed for its antiviral activities and investigate their antiviral activities against IHNV and IPNV. Activity-guided fractionation for 80% methanolic extract of Polysiphonia morrowii using a cell-based assay measuring virus-induced cytopathic effect (CPE) on cells yielded a 90% methanolic fraction, which showed the highest antiviral activity against both viruses among fractions yielded from the extract. From the fraction, two bromophenols were isolated and identified as 3-bromo-4,5-dihydroxybenzyl methyl ether (1) and 3-bromo-4,5-dihydroxybenzaldehyde (2) based on spectroscopic analyses. For both compounds, the concentrations to inhibit 50% of flounder spleen cell (FSP cell) proliferation (CC₅₀) and each viral replication (EC₅₀) were measured. In the pretreatment test, 3-bromo-4,5-dihydroxybenzyl methyl ether (1) and 3-bromo-4,5-dihy-droxybenzaldehyde (2) exhibited significant antiviral activities showing selective index values (SI = CC₅₀/EC₅₀) of 20 to 42 against both IHNV and IPNV. In direct virucidal test, 3-bromo-4,5-dihydroxybenzyl methyl ether (1) showed significant antiviral activités against both viruses while 3-bromo-4,5-dihydroxybenzaldehyde (2) was significantly effective against only IHNV. Although antiviral efficacies of both compounds against IHNV and IPNV were lower than those of ribavirin used as a positive control, our findings suggested that the red alga Polysiphonia morrowii and isolated two bromophenols may have potential as a therapeutic agent against fish viral diseases.

Streptococcus zoonotic bacteria cause serious problems in aquaculture with clinical effects on humans. A structure-antibacterial activity relationships analysis of 22 isoflavones isolated from M. tricuspidata (leaves, ripe fruits, and unripe fruits) against S. iniae revealed that prenylation of the isoflavone skeleton was an important key for their antibacterial activities (minimum inhibitory concentrations: 1.95–500 μg/mL). Through principal component analysis, characteristic prenylated isoflavones such as 6,8-diprenlygenistein (4) were identified as pivotal compounds that largely determine each part’s antibacterial activities. M. tiricuspidata ripe fruits (MTF), which showed the highest antibacterial activity among the parts tested, were optimized for high antibacterial activity and low cytotoxicity on fathead minnow cells using Box–Behnken design. Optimized extraction conditions were deduced to be 50%/80 °C/7.5 h for ethanol concentration/extraction temperature/time, and OE-MTF showed contents of 6,8-diprenlygenistein (4), 2.09% with a MIC of 40 µg/mL. These results suggest that OE-MTF and its active isoflavones have promising potential as eco-friendly antibacterial agents against streptococcosis in aquaculture.