In the course of diagnostic works of diseased ayu, Plecoglossus altivelis, and amago, Oncorhynchus rhodurus var. macrostomus, cultured in freshwater ponds in Tokushima Prefecture from 1977 to 1978, a new bacterial disease was found among them. The characteristic sign of the diseased fishes was hemorrhagic lesions of body surface, especially operculum, abdomen, and anus. A β-hemolytic streptococcus was isolated from these diseased fishes, and it proved to be able to kill ayu and amago with typical symptom of this disease by intraperitoneal injection. From the results of biochemical and biological tests, it became evident that the etiological agent of this disease was identical with the isolate from cultured yellowtail, Seriola quinqueradiata, by MINAMI et al. (1979). Yet, the species name could not be determined for the organism, because the antigen prepared from the isolate did not react with commercial antisera to Streptococcus groups A, B, C or G.
The relative efficiencies of various methods of concentration and purification of the eel virus of Europe (EVE) were examined. Whilst Freon treatment had no obvious effect on yields, EVE was effectively concentrated 10 times using polyethylene glycol (PEG), and could be concentrated 100 times using a 2 step PEG technique, or by use of high speed centrifugation as alternative to the 2nd PEG step. High speed centrifugation had the advantage of excluding UV absorptive substances other than virus, which could not be excluded by the 2 step PEG method. EVE was partially purified by sucrose linear 15-45% density gradient centrifugation at 80, 000g for 2 hrs. Well purified EVE was obtained by CsCl isopycnic centrifugation at 160, 000g for 24 hrs at a bouyant density of 1.33g/cm3. IPNV-VR299 was concentrated and purified in a similar manner to EVE and was also shown to have a density of 1.33g/cm3. The mean diameter of uranyl-acetate stained EVE was 65.5nm and that of IPNV 66.6nm. CsCl isopycnic centrifugation of EVE, but not of IPNV, revealed a faint band at a density of 1.30g/cm3 composed of many particles of average diameter 20nm apparently morphological subunits. This feature, together with the preserve of empty and collapsed particles in the main EVE band, suggested a major difference between EVE and IPNV in virus stability.