Results of a survey of Cyprinus carpio, carp, in Croatia, Yugoslavia and data from carp reared under controlled laboratory conditions, have provided new information on the enigmatic nature of Rhabdospora thelohani or “rodlet cells”. Carp larvae were obtained in the laboratory by artificial fertilization and reared so as never to be in contact with other fish. Young fish were sacrificed at 10 and 60 days of age, serially sectioned and examined for R. thelohani. In 10 day larvae, organisms were never observed. However, 60 day fry had rhabdosporans loosely associated with the afferent branchial arteries and ventral aorta distal to the bulbus arteriosus. More proximally, they appeared to aggregate at the periphery of the aorta, become associated with the endothelium while decreasing in number at the periphery and finally localized primarily inside of the bulb. Results of the survey showed one out of a total of five (20 %) 11 mm carp contained R. thelohani; the organisms were found associated with the adventitia of the ventral aorta but not the bulbar tissue per se. The bulbi of all carp >30mm in length, from all four fish farms sampled, contained varying numbers of rhabdosporans. These observations suggest a migration of rodlet cells to the bulb and suggest but do not conclusively prove that they are not parasites.
Static 96 h bioassays were conducted on milkfish fingerlings at concentrations ranging from 1.00 to 1.80mg/l KMnO4. Histopathological analyses of gills, liver, and kidney tissues revealed significant changes even in non-lethal concentrations tested. Damage became severe with in-creasing concentration and longer exposure to the chemical. Partial to complete recovery was observed in gills, liver, and kidney cells of fish exposed to KMnO4 for 96h and then maintained in KMnO4-free seawater for 240h.
This study describes attempts to establish a culture system for cells derived from tissues of the grass prawn Penaeus monodon. No cell growth was observed in cultures on fragment of gill, nerve, muscle, gut and hepatopancreas. However, an observable cell growth was obtained in the culture of gonad and heart cells. The best growth of gonad cells occurred in a culture system containing double strength (2X) Leibovitz's L-15 medium supplemented with 10% fetal calf serum (FCS), 30% muscle extract, 0.006g/ml NaCl and 10% lobster or grass prawn haemolymph. When the gonad cells were cultured in this medium for 7-10 days and incubated at 28-31°C, a confluent monolayer cell sheet was formed. The primary cell culture, predominantly of epithelioid cells, could be maintained for approximately two months at 28-31°c with a weekly change of medium. After the third consecutive subculture, the cells degenerated and dislodged from the flask surface.
In 1974, from June to July, a disease which was accompanied by a considerably high mortality occurred among juvenile colorcarp, Cyprinus carpio, at many farms in Niigata Pref., Japan. The outbreak of the disease has been observed in the same season every year since then. In most cases, a large number of fish died within a few days after onset of the disease. The characteristic symptom was swelling due to edema of the fish body. The purpose of the present study is to obtain the histopathological findings of the disease by light and electron microscopic observation. It was progressive change which induced club-shape transformation of gill lamellae by proliferation and thickening of epithelial cells of the secondary gill lamellae. The club-shape transformation began at the tip of the gill lamellae, gradually extends downward to spread all over the gill lamellae. As the degeneration proceeded, gill lamella-adhesions also began to occur. At this stage, the capillary cavities in secondary gill lamellae were almost occluded. In the gill filaments with marked pathological change electron microscopic observation revealed many virus-like particles in the epithelial cells of thickened secondary gill lamellae of diseased fish. The particle was about 250-280 nm in diameter and the profile was mulberry-like with an envelope. These morphological features suggest that this virus may be a member of the poxvirus group.
Pathogenicity of a virus isolated from ascitic fluids of diseased Hirame was studied. Twenty Hirames, average body weight 180g, were injected with 103 TCID50/fish of culture grown virus intraperitoneally and then each 10 fishes were held at 10°C or 15°C. Five Hirames were injectedwith 0.1 ml of 106 TICD50/ml intramusculally and were held at 10°C using 100l aquariums. During the 21 days post injection, 20%mortality were observed at 10°CIP and IM groups respectively but no mortality was observed at 15°C group and 10%FBS added MEM injected control groups. The virus titer of kidney tissues of died fish were 106.8-7.6 TCID50/g.
Outbreaks of motile aeromonad disease occurred among crucian carps, Carassius auratus, in game-fish ponds. Aeromonas hydrophila which produced strong β-hemolysin was isolated from the diseased fish. A histopathological study was made on the infected fish. The fish showed hemorrhage on the body surface, accumulation of red-colored ascitic fluid and enteritis with bacterial invasions. Hemosiderosis was observed in the liver, spleen and kidney. Deposition of hematoidin crystals was found in the spleen, kidney and blood vessels. Experimental infections were carried out by intraperitoneal injection with the isolate into crucian carp and resulted in manifestation of similar pathological signs to those of the naturally infected case.
The enzymatic properties of bacteriolytic substances in the homogenates of skin mucus and intestine of carp (Cyprinus carpio) was examined. The bacteriolytic activity was estimated by the turbidimetric method using acetone-ether-dried cells of Micrococcus lysodeikticus as the substrate. The skin mucus extracted with 0.001M phosphate buffer of pH 6.5 appeared the maximal bacteriolytic activity at 20°C. And intestine homogenized with 0.001M phosphate buffer of pH 6.0 showed the maximal activity at 40°C. The bacteriolytic activity of both extractions from the skin mucus and the intestine with the acidic buffer (pH 4.0) were relatively stable against to heating of 100°C for 10 minutes but those in neutral(pH 7.0) or alkaline (pH 9.0) buffer were liable to be inactivated.
Lipopolysaccharides(LPS)were prepared from different serotypes of Vibrio anguillarum, J-O-1, J-O-2 and J-O-3, and Vibrio ordalii. The LPS fractions were obtained by phenol-water extraction followed by Cetavlon treatment and high speed centrifugation. Chemical analysis of V. anguillarum J-O-1, J-O-2 and J-O-3 showed 2.5%, 4.8%and 1.3% protein;36.3%, 30.0% and 20.0% sugar and 7.1%, 11.3% and 6.6%fatty acids. Chemical analysis of V. ordalii showed 17.5%protein, 13.8% sugar and 1.4% fatty acids. The monosaccharide composition of V. anguillarum J-O-1, J-O-2 and J-O-3 was respectively:0.1%, 0.1% and0.1% 2-keto-3-deoxyhexose(KDO);3.0%, 1.9% and 0.6% pentose;32.6%, 21.6% and 12.6% hexose;10.8%, 7.6% and 2.8% 6-deoxyhexose;4.6%, 7.7% and 7.7% heptose;5.1%, 6.0% and10.2% amino sugar. The monosaccharide composition of V. ordalii LPS was:0.0% KDO, 1.5%pentose, 6.6%hexose, 4.0%6-deoxyhexose, 0.2%heptose and 3.1%amino sugar. The principal fatty acids of the V. anguillarum lipid A were:myristic acid, palmitic acid and stearic acid. The principal fatty acids constituents V. ordalii lipid A were: lauric acid, palmitic acid, stearic acidand arachidic acid.
To confirm the properties of the lipid from Edwardsiella tarda as immunosuppressor in the eel, suggested by previous studies, lipid extracted from the whole cell (Lipid), crude lipopolysaccharide (LPS), formalin killed cell (FKC) and whole cell without lipid (CWL) were injected intramuscularly in the eels. To compare the protective effect of the preparations, the eels were challenged with live E. tarda, strain EF-1. The eels immunized with the Lipid preparation did not show differences in protection against challenge in comparison with the control group, and the highest survival rate was recorded in eels immunized with the crude LPS preparation. The antibody titer of the Lipid immunization group was very low and the highest was in the crude LPS immunization group. In vitro phagocytic activity of the total blood was significantly lower in the Lipid immunization group than that found in the control group, whereas higher activity was shown in the crude LPS immunization group. The phagocytic activity of FKC and CWL immunization groups was not significantly different from that of the control group. The results confirm immunogenic properties of E. tarda lipid as immunosuppressor, which decreases the level of phagocytosis in the eel.