A novel antifungal protein (SAP) was found in the culture supernatant of a marine bacterium, Streptomyces sp. strain AP77, and was purified. This protein was characterized by chemical, biochemical, and biological analyses. By using gel filtration, the molecular mass of SAP was estimated to be 160 kDa. Structural analysis of SAP by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry suggested that SAP is composed of three heterologous protein subunits of 41.7 kDa (SAP1), 21.7 kDa (SAP2), and 18.7 kDa (SAP3) at a molar ratio of 1:1:5 (or 1:1:6). N-terminal amino acid sequence analysis and a homology search revealed that SAP1, SAP2, and SAP3 exhibit 64.3, 68.4, and 86.7% similarity to three Streptomyces coelicolor polypeptides, puromycin resistance protein (Pur8), a conserved hypothetical protein, and bacterioferritin, respectively. The MIC of purified SAP against Pythium porphyrae was determined to be 1.6 g/disk, whereas no inhibitory effect was observed at concentrations up to 100 g/disk against most of the fungal and bacterial strains tested; the only exception was relatively strong antifungal activity against Pythium ultimum (MIC, 6.3 g/disk). In vitro and in vivo toxicity tests demonstrated that SAP showed no toxicity against Porphyra yezoensis cells, human normal dermal fibroblasts, and mice at doses up to 700 g/ml (for 24 h), 250 g/ml (for 12 h), and 75 mg/kg (for 35 days), respectively. SAP was labile when it was subjected to a heated-air drying treatment, which is a great advantage in food production procedures. These results indicated that Streptomyces sp. strain AP77 might be useful as a gene source for safe transgenic Porphyra breeding for tolerance to Pythium infection.

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Four bacteria with degrading activity against the cell walls of Pythium porphyrae were successfully isolated from Porphyra‐culturing environments. The crude enzymes from these bacterial isolates, Bacillus sp. BE1, Bacillus sp. FE1, Pseudomonas sp. PE1, and Pseudomonas sp. PE2, degraded the mycelial cell walls of Pythium sp. N‐Acetylglucosamine and glucose were detected in the supernatants of Pythium cell walls treated with the enzymes from Bacillus sp. BE1, Bacillus sp. FE1, and Pseudomonas sp. PE2 as the final degrading products by high‐performance liquid chromatography, whereas only glucose was detected in the supernatant of the cell walls treated with the enzyme from Pseudomonas sp. PE1. Moreover, the activities of β‐1,3‐glucanase, β‐1,3‐1,4‐glucanase, and chitinase were observed by polysaccharolytic analysis using the enzymes from Bacillus sp. BE1, Bacillus sp. FE1, and Pseudomonas sp. PE2, whereas only the activities of β‐1,3‐glucanase and β‐1,3‐1,4‐glucanase were found during analysis using Pseudomonas sp. PE1. β‐1,4‐Glucanase, β‐1,6‐glucanase, and mannanase activities were not detected in any of the crude enzymes obtained from these isolates. From the four isolates, the molecular weights (MW) of β‐1,3‐glucanase and chitinase were estimated to be approximately 50 000–100 000 by the ultrafiltration method. Two Pseudomonas spp. were also suggested to have β‐1,3‐1,4‐glucanases with MW of 50 000–100 000. However, the MW of β‐1,3‐1,4‐glucanases from the two Bacillus spp. might be close to 50 000 or they produce at least two β‐1,3‐1,4‐glucanases with MW of 30 000–50 000 and 50 000–100 000, respectively.

We attempted to screen for telomerase inhibitingactivity in vitro from a total of 304 marine algae samples which were collected from various Japancoasts by nonradioisotope telomeric repeatamplification protocol (Non-RI TRAP) assay using humanleukemia MOLT-4 cells. Ten of the MeOH extracts andtwo of the PBS extracts from those algal samplesshowed telomerase inhibiting activity. In particular,the MeOH extract from a green alga, Caulerpasertularioides strongly inhibited telomerase activitywhen added to MOLT-4 cell culture at a level of 1.25%(v/v), suggesting that it may be possible to developa novel anti-cancer agent in view of its specificanti-telomeric property.

A simple, low-cost and efficient bioreactor system for production of anti-infectious hematopoietic necrosis virus (IHNV) substances by bacteria isolated from aquatic environments was devised on a laboratory scale. Whole living cells of anti-IHNV substances producing bacteria, Pseudomonas sp. 51BBW-29 and Alteromonas sp. 48HS-27, were immobilized in calcium alginate beads prepared by 1% sodium alginate, 0.45M CaCl2, and C2Y or MC2Y broth. Eighteen g of the beads were packed into a 25 ml reaction column and incubated at 15℃ for 5 days in a circulating broth medium at a flow rate of 0.35ml/h. After activation of the immobilized bacteria, the beads were incubated in a continuous flow of fresh broth with intermittent exposure to air. In the beads incubated under these conditions, the immobilized bacteria grew well at the gel surface and high viability of approximately 10^10 CFU/bead was retained for at least 6 days from day 8 to 14. During 31 days of the experiment with the bioreactor system, the highest production was 69.3 ⊿IHD50 for Pseudomonas sp. 51BBW-29 (at 14-17 days incubation) and 48.0 ⊿IHD50 for Alteromonas sp. 48HS-27 (at 8-11 days incubation), which were 2.5 and 2.7 times higher than those of conventional batch culture (27.3 and 18.0 ⊿IHD50 for the two strains at 2-5 days incubation). The immobilized bacteria continuously produced high levels of anti-IHNV substances for 15 days (day 5-20)

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