The prophylactic effects of chemicals and immunostimulants on theTetrahymena infection of guppies (Poecilia reticulata) were studied through experimental infections.Tetrahymena pyriformiswas used in an experimental infection on a guppy population pre-treated by the following method; the base of the caudal fin was covered with a 10% acetic acid-soaked cotton strip for 3 min. The study consisted of screening anti-Tetrahymenachemicals, testing prophylactic effects of selected chemicals or immunostimulants, and finally evaluating the combination effects of the chemicals and immunostimulants. The single uses of chemicals or immunostimulants did not completely prevent theTetrahymenainfection. Prevention was attained by the combination of a 0.5% sodium chloride bath and a feeding of C-UPIII (a Chinese herb mix) diet.
An in situ hybridization (ISH) using digoxigenin-labeled 16S rDNA probes was developed to detect the causative agent of bacterial coldwater disease, Flavobacterium psychrophilum. Strong, specific hybridization was observed for all F. psychrophilum strains tested and no crosshybridization was seen for 13 other bacterial species. Distribution of F. psychrophilum in various organs of experimentally and naturally infected ayu was investigated by the ISH. A group of the experimental fish were given a sting at the skin and then immersed in a suspension of F. psychrophilum (106CFU/mL) for 1 h at 15°C and another group were subcutaneously injected with F. psychrophilum at a dose of 1.5×107 CFU/fish. In both experiments, F. psychrophilum was detected in the muscle, gills, heart, kidney and spleen, but not in the brain, liver, stomach, intestine, or pyloric caeca during the experiment period (7 h). A large number of F. psychrophilum were found at the stung or injected sites of the muscle, while no bacteria were detected at the intact muscle. In some of naturally infected ayu collected in fish farms, F. psychrophilum was detected in the liver, pancreas, stomach, pyloric caeca and intestine. The muscle of the naturally infected ayu was severely infected with F. psychrophilum, but no bacterium was detected in the brain.
Challenges of the Japanese flounder Paralichthys olivaceus with the monogenean Neoheterobothrium hirame were carried out to clarify the cause of the anemia, recently prevailing in the Japanese flounder in Japan. Hematological changes very similar to those previously reported in both wild and cultured anemic flounder were produced by the challenges with H. hirame. In the challenged flounder, the hemoglobin concentration decreased considerably and abnormal immature and mature erythrocytes having vacuolated or weakly stained cytoplasm, which characterize this anemia, appeared. The relationships among hemoglobin concentration, the ratio of immature erythrocytes and the above morphological changes in the blood smear of the challenged fish were very similar to those in the naturally affected anemic flounder. These hematological changes showed a clear correlation with the number of adult worms. These results, together with the previous findings, suggest that N. hirame is the main cause of the anemia prevailing in the Japanese flounder.
A survey of Neoheterobothrium hirame infection of Japanese flounder Paralichthys olivaceus in several places in Japan (Kyoto, Fukui and Ibaraki) was conducted to examine the relationship between the development of the parasite and its microhabitats. The parasite first attached to the gill filaments, then migrated to the gill arches/rakers and finally moved to the buccal cavity wall. This migration was associated with growth and maturity of the parasite. Some worms already reached maturity on the gill rakers, but produced eggs only on the buccal cavity wall. The histology of infected tissues was examined to determine the mode of attachment and pathological effects of the parasite. On the gill filaments, the parasite used, at first, marginal hooks and hamuli to attach to the proximal region of the secondary gill lamellae and later, clamps to grasp these lamellae. On the gill arches/rakers, clamps were used to attach to the epithelium. On the buccal cavity wall, clamps were used initially for attachment to the tissue surface but later the haptor penetrated the underlying connective and muscle tissues until the haptor and almost the entire isthmus region were embedded deeply in the host tissue. Only a mild inflammatory response was observed on the gill filaments and in the epithelium of the gill arches/rakers, respectively, whereas a strong host response and necrosis were associated with the prolonged attachment of the parasite to the buccal cavity wall.
Neoheterobothrium hirame infection of 0-year wild Japanese flounder Paralichthys olivaceus was investigated. This study was based on annual samples (preserved in formalin or alcohol) totalling 316 fish caught off Igarashi-hama in July and/or August of 1989-1993 and 505 fish caught off Murakami City in August of 1993-1999, Niigata Prefecture. A stirring method was developed to collect the monogenean from the fixed gill filaments. All the parasites were dislodged from the fixed gill filaments by stirring gills of individual hosts in 150 mL of water with a magnet (40 mm long and 8 mm in diameter) at 1150-1200 rpm for 20 min and 30 min for the 0-year fish and 1-year fish, respectively. Adult parasites were macroscopically observed on the buccal cavity wall, whereas immature ones on the gill arches and rakers were detected under a stereomicroscope. No N. hirame was found from 1989 to 1992. The earliest records were from samples caught off Igarashi-hama and Murakami City in 1993, though the prevalence of infection was very low. The parasite occurrence changed annually in the Murakami samples. The annual fluctuations may have been influenced by the infection level of co-existing 1-year fish and the population size of 0-year fish recruited each year. It is inconclusive whether or not the monogenean is an indigenous parasite of Japanese flounder in this area.
The effect of low incubation temperature on the embryonic development and larval hatching of the monogenean Diplectanum aequans, a gill parasite of sea bass Dicentrarchus Iabrax, was studied. Groups of eggs of D. aequans were incubated at 5°C for different time periods (from 24 h to 28 days) before incubation at room temperature (18°C). Percentages of hatched larvae, aborted larvae and undeveloped embryos were estimated in comparison with the control group. Results show that eggs of the parasite are able to survive at 5°C, with a percentage that decreases as the incubation time at 5°C increases, reducing the rate of larval hatching.
An outbreak of a disease occurred in hybrid striped bass (20-25 cm in length) Morone saxatilis × Morone chrysops cultured in Taiwan. No significant external sign was found, whereas prominent nodules were observed in internal organs, such as the spleen and kidney. The most severe histopathological changes were necrotic foci and multiple granulomas in haematopoietic tissues of the spleen and kidney. The results of biochemical examination and PCR-based detection indicate that the disease was caused by Photobacterium damselae subsp. piscicida. This is the first reported outbreak of pasteurelosis in cultured hybrid striped bass.
Susceptibilities of spangled emperor Lethrinus nebulosus, estuary cod Epinephelus malabaricus and cobia Rachycentron canadum cultured in subtropical Japan to red sea bream iridovirus (RSIV) were studied by experimental infections. Estuary cod showed as high susceptibility to RSIV as red sea bream Pagrus major. In contrast, the susceptibilities of spangled emperor and cobia to the virus were comparatively low. The result corresponds with the occurrence of the disease in the net cages.
The capability of Streptococcus iniae to cause disease in Japanese flounder was investigated by oral and bath (30 min) challenges with low, medium and high inoculation doses. Only with the highest inoculation dose, 9.9×107 CFU/100 g body weight, the oral challenge induced deaths. In the bath challenge deaths were induced even at the lowest inoculation dose, 2.9×103 CFU/mL water. Hemorrhagic lesions on the fins were observed in dead fish challenged by both methods. It is suspected that S. iniae entered through the body surface such as abrasive sites of the fins to cause disease.