Pike eels Muraenesox cinereus (41-116 cm in total length; n=23) purchased from fish markets in Aichi Prefecture were infected with the digenetic trematode Aerobiotrema muraenesocis in the swim bladder (11 out of 23 fish; intensity of infection 1-27). Effects of the infection on the host were studied and partial life history of this trematode was postulated. Eggs of the trematode were spherical or sub-spherical. They could hatch in the water with a wide range of salinity (1/3 to undiluted). Hatched larvae had an unusual appearance, with long cilia limited on the antero-lateral sides of their body. Histochemical staining revealed that the trematode sucks blood from the swim bladder of the host. Moreover, severe histopathological changes were observed in the tunica interna of the swim bladder, including deep and irregular undulations of the epithelium, marked thickening of the tunica propria and hemorrhaging ulcers within the tunica interna. In case where these pathological changes had proceeded, the intimate association of the epithelium with the capillaries in the tunica propria was severely disturbed. According to these histopathological observations, it appears that heavy infection with this trematode may cause lowered functions of the host's swim bladder.
The rhabdoviruses isolated from EUS-affected snakeheads in the Philippines was tested for pathogenicity to healthy, naive snakehead Ophicephalus striatus fry, fingerlings and juveniles. Virus exposure of naive snakehead fry and fingerlings by bath at 20-22.5°C resulted in significant mortalities (p<0.01) with no apparent lesions. Naive snakehead juveniles when injected intramuscularly (IM) with the EUS-associated rhabdovirus at ambient water temperature (28-32°C) did not develop any lesion. However, similarly treated snakehead juveniles at 20-22.5°C developed dermal lesions 3-5 days following treatment. The lesions progressed from slight to moderately advanced lesions by days 10-12 but not to deep ulcers as exhibited by naturally EUS-affected snakeheads. Mean mortalities were higher in the virus-injected fish (72%) compared to those in controls (33%). Moreover, the virus was reisolated from fish in the 20-22.5°C treatment but not from fish in the 28-32°C treatment. Virus from infected tissue filtrate and the virus passaged 3 or 4 times induced similar dermal lesions if the rhabdovirus concentration was 103 TCID50/fish or higher. When administered orally, by bath, by intraperitoneal (IP), IM and subdermal injections to snakehead juveniles, only the latter two viral routes induced dermal lesions. However, IP injection of the rhabdovirus caused 75% mortalities but none in control fish. The results demonstrate the pathogenicity of the rhabdovirus isolates to naive snakeheads at low (20-22.5°C) rearing water temperatures.
Hirame rhabdovirus (HIRRV) was first reported from flounder (Paralichthys olivaceus) in Japan, and has recently been isolated from an aquaculture farm in the Tongyoung area of Korea. Total RNA isolated from RTG-2 cells infected with HIRRV was used in RT-PCR to obtain a cDNA coding for the glycoprotein of HIRRV Korean strain, CA-9703. Around half sized G protein in the C-terminal region was expressed as glutathione-S-transferase fusion protein, and used as viral vaccine against HIRRV. Flounder fry were vaccinated with low (50μg/mL) and high (300μg/mL) doses of vaccine by immersing them for 7 min in vaccine solution. After 1 month, the fish were challenged with HIRRV either by immersion or intraperitoneal injection. Specific losses of fish treated with low and high vaccine doses by intraperitoneal injection challenge were both 6.7%, in contrast to 43.3% in non-vaccinated fish. Losses of vaccinated fish by immersion challenge were 1.7% and 0% for low and high vaccine doses, respectively, and 15% for the non-vaccinated control.
A study was conducted to investigate the antifungal activity of some bacteria isolated from the lesions of salmonid fishes with saprolegniasis. Five of 47 isolates inhibited the growth ofSaprolegniaparasitica H2 andS. salmonisNJM 9851 on solid media. Among 8 media examined, BHI, HI, and to a lesser extent NA were suitable for demonstrating the antifungal activity. The inhibitory activity against the testedSaprolegniadid not reside in supernatant of bacterial broth culture of the selected strains. The strains which produce antifungal substances belong to the generaAlteromonas, Pseudomonas, andAeromonas. These results suggest that the selected bacteria are natural competitors ofSaprolegia, and may be used for biocontrol ofSaprolegniain aquaculture.
The effect of high pH (pH 9.25) of rearing water on the prevention of infection of the brine shrimpArtemia salinalarvae with four fungal species of Lagenidiales was examined. The four fungi, all pathogenic to crustaceans, wereHalocrusticida parasiticaisolated from greasyback shrimpMetapenaeus ensis, Haliphthoros sp. andHaliphthoros milfordensisisolated from swimming crabPortunus trituberculatusandLagenidium callinectesisolated from mud crabScylla serrata. Thein vitrogrowth ofHalocrusticida parasitica was reduced at pH 9.25 and no infection was observed in the brine shrimp exposed to zoospores in the sea water at pH 9.25. Thein vitrogrowth ofHaliphthoros sp. andHaliphthoros milfordensiswas reduced at pH 9.25 and infection rates at pH 9.25 were about half of the rate at pH 8. The growth ofL.callinecteswas not arrested at pH 9.25 and the infection rate at pH 9.25 was 100%. These results indicate that the pH adjustment of rearing water to 9.25 can be used to prevent infections withHalocrusticida parasiticaand the two species ofHaliphthorosin swimming crab larvae, but not for the infection withL. callinectes.
Seasonal occurrences of the abnormally enlarged ovary of Pacific oystersCrassostrea gigascaused by an unidentified intracellular protozoan parasite was investigated in Gokasho Bay, Mie Prefecture, Japan. Cultured and wild oysters were collected bimonthly for one year from July 1996, and seasonal fluctuations in the prevalence of infection, parasite development and histological changes of the infected tissue were examined. Parasites, very similar toMarteilioides chungmuensisinfecting Pacific oysters in Korea, had 8 forms, 4 of which were frequently observed, while the other 4 forms, which were not described inM. chungmuensis, were rare. The total prevalence of infection (percentage of infected oysters positive for at least one of the two examinations : gross and smear inspections of the ovary) was usually higher than the gross prevalence of infection, indicating that infected oysters included ones without visible legions. In cultured oysters, the prevalence of infection increased in summer, suggesting that active multiplication of the parasite takes place in warm water months. From autumn through spring, no substantial change in the total prevalence of infection was observed in cultured oysters (18-20%), while it continued to decline in wild oysters during that period. These differences between cultured and wild oysters could be attributed to the oyster size and environmental factors such as salinity. Histologically, the number of infected oocytes increased in summer, but decreased in winter. From the variations in the parasite morphology and maturation of oocytes, the life cycle of the parasite and the mechanism of the legion formation are postulated.
The presence of white spot syndrome virus (WSSV) in wildPenaeus monodon, other wild shrimps, and in non-cultured crustaceans from shrimp ponds/ghers in Bangladesh was studied by polymerase chain reaction (PCR).Wild shrimps viz.P. mondon, P.semisulcatus, P.indicus, Metapenaeus monoceros, M. brevicornisandPalaemon styliferuswere positive for WSSV.The non-cultured shrimps M. monoceros, M. brevicornis, freshwater prawnMacrobrachium rosenbergii, crabsScylla serrataandPseudograpsus intermediusfrom shrimp ponds/ghers were WSSV positive.This is the first study on the detection of WSSV in Bangladesh by PCR and first report ofP.styliferus, M.monoceros, M. brevicornisandP.intermediusas WSSV carriers.
A simple assay to determine the proliferation of larval and juvehile fish splenocytes using Alamar Blue (AB) was studied. Splenocytes were separated by Percoll gradient from Japanese flounderParalichthys olivaceus (1-year-old), then the proliferation of splenocytes stimulated by mitogen (concanavalin A : ConA, pokeweed mitogen : PWM, or lipopolysaccharide : LPS) was detected using AB. The relationship between number of splenocytes and specific absorbance exhibited a positive significant correlation. The optimum condition of this assay was 5×105 cells/well of splenocytes with mitogens (ConA 100μg/mL, PWM 10μg/mL or LPS 1μg/mL) for 72 h. Proliferation of each splenocyte from juveniles of Japanese flounder and larvae of Japanese parrotfish Ophegnathus fasciatus and tiger puffer Takifugu rubripes was detectable by this assay.
Pathogenicity of a marine birnavirus (AY-98) isolated from diseased ayu Plecoglossus altivelis was investigated. Cumulative mortalities in the intraperitoneally injected groups of ayu in two weeks were 70% (dose 105.4 TCID50/fish) -71% (105.8 TCID50/fish) in duplicated experiments. Histologically, the naturally and experimentally infected fish showed brain congestion and pancreatic necrosis. From the symptoms of body deformity and congestion in the brain, it was thought that AY-98 is similar to viral deformity virus, the birnavirus isolated from yellow tail Seriala quinqueradiata with deformity.
A symposium entitled “Current status of fish vaccine development in Japan” was held by the Japanese Society of Fish Pathology in Tokyo on 29th March, 2001. This was aimed to have an overview of the status quo of fish vaccine developments in Japan, especially those for marine fishes. In Japan, a vibrio vaccine to ayu Plecoglossus altivelis was first developed in 1988 and five vaccines have been totally developed until now. In this symposium, ten subjects were included in five categories : (1) Vaccine-use system, utilization and practical problems, (2) Vaccines developed or being developed, (3) Genetic engineering techniques and adjuvants for vaccines (4) General control methods to prevent fish diseases, and (5) Problems confronting the development of fish vaccines in Japan. Vaccines will contribute to make safe products in aquaculture, which can give “peace of mind” to consumers. Vaccines developed in Japan have been effective to protect fish from the target diseases and it was emphasized that fish condition should be kept healthy to get a good result by vaccination.