Chloramphenicol (CM)- and tetracycline (TC)-resistant bacteria which could grow on Heart Infusion agar containing 6.2 mcg/ml of CM or TC, respectively, were isolated with high frequencies from the intestinal tracts of cultured Ayu (Plecoglossus altivelis) and the water of Ayu culturing ponds in Tokushima Pref. CM- and TC-resistant bacteria were not isolated from the intestinal tracts of wild Ayu collected from the river Nakagawa in Tokushima Pref. The principal drug resistant bacteria isolated from the intestinal tracts were Aeromonas hydrophila and Vibrio anguillarum. Drug resistant Pseudomonas, Enterobacter aerogenes, E. cloacae, Escherichia coli, Hafnia, Salmonella arizonae and other unidentified Enterobacteriaceae were occasionally found. Drug resistant bacteria isolated from the water of ponds were common to those from the intestinal tracts. However, drug resistant E. coli, Psudomonas and V. anguillarum were not isolated from the water. All drug resistant V. anguillarum strains carried R plasmids. R plasmids were detected in a part of the Hafnia, E. cloacae, A. hydrophila, unidentified Enterobacteriaceae and E. aerogenes strains. The drug resistance markers present in the R plasmid bearing strains were in most cases sulfonamides streptomycin CM and TC.
The pharmacokinetic and residual examinations of erythromycin(EM)were studied in cultured yellowtails Seriola quinqueradiata orally administered a single or multiple dose (s) of 50mg/kg/day with the following results. Orally administered EM produced peak levels in blood and tissue 1 to 3 hr post dosing and the levels in liver, kidney and spleen were about 4 times as high as that in blood.The level in muscle was approximately equal to that in blood. A single dose of EM produced blood and tissue levels which were 10 to 100 times as high as the MIC for Streptococcus sp.1 hr post dosing, and higher levels than the MIC were detected 24 hr following administration. Blood and tissue levels of EM after 10 consecutive administrations were almost same or slightly higher than those obtained by the single administration, though their distribution patterns were similar. After the withdrawal of 10 consecutive dosing, levels of EM in blood and tissue decreased exponentially, and biological half-lives were blood;5.00-7.78hr, liver;8.37-9.75hr, kidney;14.80-15.63hr, spleen;14.32-15.89 hr, muscle;7.15-8.17hr. A level below the assay limits(0.03-0.08μg/ml or g)was reached 144-168 hr post dosing. No evidence of long term residue was found in any of the tissues tested.
Efficacy of erythromycin (EM) against spontaneous streptococcal infection in cultured yellowtails Seriola quinqueradiata was evaluated clinically. The present studies given seven experiments were accomplished from August to October in 1978. The fish tested were 1st (80, 500 fish) and 2nd (29, 100 fish) year fish of cultured yellowtails being raised at fish farms in Kochi and Kagoshima prefectures. All of the clinical isolates of Streptococcus sp. from naturally infected fish were susceptible to EM, tetracycline, ampicillin and chloramphenicol, but resistant to sulfamonomethoxine. The minimal inhibitory concentration of EM was ranged from 0.1 to 0.2μg/ml and was the lowest in the tested drugs. In the clinical studies, EM was mixed in minced mackerel or mackerel pikes with formula feed and a binder, and consecutively administered for 4-7 days at the dose of 25-50mg/kg/day. The diseased fish, in every experiment, responded to EM with reduction in mortality and were found to be controlled of the disease during medication or within a few days post dosing. The efficacy of EM against streptococcal infection was more remarkable than those of oxytetracycline (50-80mg/kg/day) and ampicillin (20mg/kg/day). EM was also effective against the disease which couldn't cured by ampicillin. The administration of EM caused no clinical abnormality in about 100, 000 fish of cultured yellowtails. The results of the present study suggest that EM can be used for the control of streptococcal infection in cultured yellowtails.
The phagocytic system in goldfish was studied through intraperitoneal injection with India ink. Fish were killed at intervals ranging from 30 min to 50 days after injection. Histological research was performed on spleen, kidney, heart, liver, gills and blood. On the first day after injection, the spleen and kidney became black. Phagocytosis of carbon particles was performed from the first day after injection by the sheath cells in the sheathed artery of the spleen, by the reticuloendothelial cells throughout the haematopoietic tissue of the kidney, and by the endothelial cells in the ventricle of the heart. Carbon phagocytizing macrophages in the kidney and spleen formed aggregates within pre-existing aggregates of melano-macrophage on the fourth day after injection. Those aggregates were observed for 50 days. There was no phagocytosis of carbon particles observed in the liver and gills. Two types of mononuclear phagocytes phagocytizing carbon particles were observed in smears of blood. The large phagocytes measured between 14 and 16μ in diameter and the nucleus occupied almost half of the cell. The small phagocytes measured between 6 and 7μ in diameter and the nucleus occupied almost all of the cell. Although the small phagocytes were observed in smears during the period from the first day to the thirty-sixth day after injection, the large ones were only observed from the first day to the seventh day. The significance of the phagocytic system in fish is discussed.
Antibacterial activity of furazolidone (N-(5-nitro-2-furyliden)-3-amino-2-oxazolidone, FZD) against Edwardsiella tarda isolated from diseased eel, Anguilla japonica, and therapeutic effect of the drug by medical bath against experimental Edwardsiellosis in young eel were studied. Minimal inhibitory concentration of FZD against 30 strains of E.tarda was 0.03-5.0ppm and sensitivity distribution curve had two peaks. One peak was at 0.07 ppm and the other at 1.2 ppm. FZD was highly effective by medical bath at over 1ppm against experimental Edwadsiellosis with E. tarda A strain (MIC was 0.07 ppm) and effective at over 4 ppm with E. tarda B strain (MIC was 1.2 ppm).
Examinations were made on the pesticidal effects of trichlorfon against Argulus coregoni parasitic on yearling yamame (Oncorhynchus masou) and on the lethal effects of trichlorfon on four species of juvenile salmonids such as yamame, brook trout (Salvelinus fontinalis), rainbow trout (Salmo gairdneri) and brown trout (Salmo trutta). The pesticidal effects of trichlorfon causing 100% mortality of A. coregoni at 20°C and pH 7.0 were obtained by the immersions in 6.25 ppm for 1 hr, 12.5-50ppm for 30 min, 100 ppm for 10 min. The higher temperature and the higher pH value were found to increase the effects. The immersion of the following concentrations and durations removed more than 90% of the parasite from the host : 12.5 ppm for 60 min, 40 ppm for 30 min, 100 ppm for 20 min, 200 ppm for 10 min and 400 ppm for 30 min at 17-18°C, although the immersion in 3.13 ppm for 60 min, 10 ppm for 30 min and 80 ppm for 5min showed poor effects of less than 10% removal. As to the lethal effects on salmonids, the immersion in the solution up to 800 ppm for 1 hr at 16-17°C caused neither death nor abnormal behavior of each 10 test fish. However, the high mortality occurred in the immersion for longer period up to 18 hrs in the solutions ranging 6.25 to 400 ppm at 16-18°C. Although there were slight differences in the susceptibility among the four species, S. salvelinus was relatively susceptible while S. gairdneri being resistant to trichlorfon.
Laboratory and field experiments were made to examine the effects of time of day, water depth, and roughness of substrata on the egg deposition of Argulus coregoni. Egg-laying occurred normally at night (98%, 93/95) ; however, mature females laid eggs even in the daytime under experimentally dark conditions. The oviposition was thus considered to be influenced by the absence of light. The relationship between water depth and egg deposition was examined in trout ponds of 65 cm and 140 cm deep by hanging acrylic plates as substrata for the egg deposition, and the clusters of eggs deposited on the plates were analyzed. The clusters were found to be more abundant nearer the bottom of both ponds : 88% of the total number of clusters deposited at a depth of 45-65 cm in the pond of 65 cm depth and 74% at a depth of 100-140 cm in the 140 cm-deep pond. The parasite showed a preference of roughness of substrata. Only a few egg clusters were deposited on transparent glass (smooth substratum of less than 0.5 μm maximum height of roughness) and on a wooden plate (rough substratum of more than 50 μm), whereas abundant egg clusters were found on frosted glass or unglazed tile with a roughness of about 9 μm. Methods of control of A. coregoni utilizing these results are also discussed.
Epizootics of edwardsiellosis has been frequently occurring among pond-cultured eels, Anguilla japonica in Japan. Accurate diagnosis of the disease by means of conventional cultural method requires a day or more for completion. In this report, basic investigations for the application of direct immunofluorescence as rapid diagnostic tools of the disease were made. Antiserum was obtained from rabbit immunized with formalin-killed E. tarda (strain E-8) isolated from eel. The titer against formalin-killed homologous antigen was 1: 2048 by agglutination test. IgG of the antiserum was fractionated by ammonium sulfate precipitation and conjugated with FITC. The F/P molar ratio of labeled antibody was determined as 1.3. The labeled antibody had staining titer of 1:16 or 1:32 against 18 strains of E. tarda, including homologous strain E-8, isolated from pond-cultured eels at various regions in Japan. But no crossreaction was observed when 2 strains of V. anguillarum, 3 of Vibrio sp., 3 of A. liquefaciens, 8 of Aeromonas sp., one of Ps. anguilliseptica, one of Ps. fluorescens, one of Ps. putida, one of Escherichia coli, 2 of Salmonella sp., one of S. marcescens and one of F. columnaris were stained. The specific fluorescence of target antigen (E. tarda) was observed when acetone-fixed impression preparations of liver, kidney, spleen and injection site of muscle derived from artificially infected eel were stained with 4-fold dilution of labeled antibody. The specificity of the reaction on impression preparation was ascertained by one-step inhibition test. Stainability of E. tarda on impression preparation was stable for at least 11 days at 37°C. Stainability of E. tarda on impression preparation of liver tissue that had been preserved in 10% formalin for at least 7 days, though became slightly weaker, was not disappeared.