Ilhan Altinok

Histopathology of Rainbow Trout Exposed to Sublethal Concentrations of Methiocarb or Endosulfan

Liver, spleen, trunk kidney, gills, and brain of rainbow trout (Oncorhynchus mykiss) were examined histologically after exposure to different concentrations of methiocarb (2.5 and 3.75 mg/L) or endosulfan (0.6 and 1.3 μg/L) for 21 days. Histological recovery was also studied by maintaining the pesticide-exposed fish in a freshwater system for an additional 30 d. Lesions were not evident in liver, kidney, spleen, or brain of fish exposed to either concentration of methiocarb for 21 d. Lesions were observed in gills, liver, spleen, and trunk kidney (but not brain) of rainbow trout exposed to either concentration of endosulfan. There was no concentration-related effect observed on the histopathological lesions. After 30 days of recovery, fish had no histological lesions in gills, kidney, spleen, liver, or brain. Therefore all the changes observed during exposure were reversible.

Development of multiplex PCR assay for simultaneous detection of five bacterial fish pathogens.

A multiplex polymerase chain reaction (PCR) method was designed for the simultaneous detection of the five major fish pathogens, Aeromonas hydrophila, Aeromonas salmonicida subsp. salmonicida, Flavobacterium columnare, Renibacterium salmoninarum, and Yersinia ruckeri. Each of the five pairs of oligonucleotide primers exclusively amplified the targeted gene of the specific microorganism. The detection limits of the multiplex PCR was in the range of 2, 1, 1, 3, and 1CFU for A. hydrophila, A. salmonicida, F. columnare, R. salmoninarum, and Y. ruckeri, respectively. Multiplex PCR did not produce any nonspecific amplification products when tested against 23 related species of bacteria. The multiplex PCR assay was useful for the detection of the bacteria in naturally infected fish. This assay is a sensitive and specific and reproducible diagnostic tool for the simultaneous detection of five pathogenic bacteria that cause disease in fish. Therefore, it could be a useful alternative to the conventional culture based method.

Assessment of acute toxicity and histopathology of the fungicide captan in rainbow trout.

Acute toxicity of the fungicide, captan, to juvenile rainbow trout was evaluated under static-renewal test condition. Actual concentrations of captan ranged from 0.05 to 1.00 mg/L. The concentrations of captan that killed 50% of the rainbow trout (3.11±0.8 g) within 24 (24 h; LC(50)), 48, 72 and 96 h were 0.57±0.09, 0.49±0.10, 0.44±0.11 and 0.38±0.13 mg/L (95% confidence limits), respectively. None of the unexposed control fish died and the first fish died 6 h after exposure to captan (≥0.65 mg/L). Hypertrophy, separation of epithelium from lamellae, lamellar fusion, and epithelial cell necrosis were observed on captan exposed fish. Gills also had scattered areas of focal lamellar hyperplasia. Fish exposed to fungicide had inflammation and necrosis in liver, trunk kidney and spleen. In order, the most affected organs were gill, trunk kidney and liver.

Effects of water quality and fish size on toxicity of methiocarb, a carbamate pesticide, to rainbow trout

The acute toxicity of methiocarb in juvenile rainbow trout (Oncorhynchus mykiss, 3.25±0.79g) was evaluated in glass aquaria under static conditions. Nominal concentrations of methiocarb in the toxicity test ranged from 1.25 to 7.50mgL(-1). The concentrations of methiocarb that killed 50% of the rainbow trout within 24-h (24-h LC(50)), 48-h LC(50), 72-h LC(50), and 96-h LC(50) were 5.43±0.19, 5.04±0.18, 4.95±0.19, and 4.82±0.21mgL(-1) (95% confidence limits), respectively. Mortality of fish increased with increasing water temperature. Increasing alkalinity from 19mgL(-1) as CaCO(3) to 40, 60, or 90mgL(-1) as CaCO(3) significantly decreased mortality of fish. Total hardness ranging from 50mgL(-1) as CaCO(3) to 147mgL(-1) as CaCO(3) did not affect mortality of fish exposed to methiocarb. Fish exposed to methiocarb had histological alterations such as lamellar edema, separation of epidermis from lamellae, and lamellar fusion. Methiocarb exposed fish had necrosis between molecular and granular layer of cerebellum where Purkinje cells present. Results indicate that alkalinity, temperature, and fish size affect methiocarb toxicity of rainbow trout.

Histopathological changes induced by maneb and carbaryl on some tissues of rainbow trout, Oncorhynchus mykiss

Acute toxicity of the pesticides, maneb and carbaryl, to juvenile rainbow trout were evaluated under static-renewal test conditions. Actual concentrations of maneb ranged from 0.10mg/L to 2.00mg/L and carbaryl ranged from 0.20mg/L to 3.90mg/L. The concentrations of maneb that killed 50% of the rainbow trout (3.27+/-0.9g) within 24-h (24-h; LC(50)), 48-h, 72-h and 96-h were 1.19+/-0.12, 1.04+/-0.11, 0.92+/-0.12 and 0.81+/-0.14mg/L (95% confidence limits), respectively. LC(50) values of carbaryl for 24-h, 48-h, 72-h and 96-h were 2.52+/-0.71, 2.16+/-0.63, 1.71+/-0.46 and 1.39+/-0.15mg/L, respectively. None of the unexposed control fish died and the first fish died 6h after exposure to maneb (>or=1.30mg/L), and carbaryl (>or=2.60mg/L). Lamellar edema, separation of epithelium from lamellae, lamellar fusion, swelling of the epithelial cells and epithelial cell necrosis were observed on maneb and carbaryl exposed fish. Gills also had scattered areas of focal lamellar hyperplasia. Fish exposed to pesticides had inflammation and focal necrosis in liver, trunk kidney and spleen. Maneb and carbaryl had similar histopathological lesions. In order, the most affected organs were gill, trunk kidney and liver.

Histopathological changes in rainbow trout (Oncorhynchus mykiss) after exposure to sublethal composite nitrogen fertilizers.

Subchronic toxicity of composite inorganic fertilizers such as ammonium sulfate [fertilizer A; (NH(4))(2)SO(4); 21% NH(4)-N)], composite fertilizer 15-15-15 (fertilizer B; commercial formulation: 15% NH(4)-N, 15% phosphorus, and 15% potassium oxide), and composite fertilizer 25-5-10 (fertilizer C; commercial formulation: 25% NH(4)-N, 5% phosphorus, and 10% potassium oxide) on the skin, liver, kidney, pancreas, and gills of the juvenile rainbow trout (Oncorhynchus mykiss) was studied in two-week toxicity tests under static-renewal test conditions. Fish exposed to sublethal concentrations of fertilizers did not show any behavioral abnormality compared to control groups. Histological lesions were observed in skin, gills, liver, pancreas, and trunk kidney of the fish. In the epidermis, degenerated/vacuolated epithelial cells, microcystic dilatations, and intracellular edema of mucus cell were observed. Liver had swollen and degenerated hepatocytes without losing adenoid structure. Hematopoietic tissues had necrosis and vacuolar degeneration on proximal tubules of the kidney. In order, the most affected organs were skin, liver, and kidney.

Effects of some pesticides on the vital organs of juvenile rainbow trout (Oncorhynchus mykiss)

Gill, trunk kidney, spleen, and liver of rainbow trout (Oncorhynchus mykiss) were examined after exposure to different sublethal concentrations of carbosulfan (25, 50 and 200 μgL(-1)), propineb (3, 6 and 24 mgL(-1)), and benomyl (2, 5 and 20 mgL(-1)) for 14 days. Lesions were observed in gill, trunk kidney, spleen, and liver of rainbow trout exposed to either concentration of pesticides. The most important lesions were determined in the highest concentrations of pesticides. Lamellar fusion, lamellar hyperplasia, epithelial lifting, vacuolization of epithelial tissue, epithelial necrosis, hypertrophy and sloughing of epithelium were observed on fish exposed to carbosulfan, propineb and benomyl. Fish had cell necrosis, degeneration and oedemas in liver, trunk kidney and spleen. None of these lesions were seen in control fish.

Multiplex PCR assay for detection of four major bacterial pathogens causing rainbow trout disease

A multiplex PCR (mPCR) method was designed for the simultaneous detection of 4 major fish pathogens, Flavobacterium psychrophilum, Lactococcus garvieae, Pseudomonas aeruginosa, and P. putida. Each of the 4 pairs of oligonucleotide primers exclusively amplified the 16S rDNA gene of their targeted microorganism. The average detection limits for each organism amplified by mPCR were 2 colony-forming units (CFU) of F. psychrophilum, 3 CFU of L. garvieae, 3 CFU of P. aeruginosa, and 5 CFU of P. putida in mixed cultures. Multiplex PCR did not produce any nonspecific amplification products when tested against 28 related species of bacteria. High amounts of DNA from 1 bacterial species had a significant effect on the amplification sensitivity of the other bacterial species when these were present in lower concentrations in the multiplex reaction. The mPCR assay proved useful for the detection of the bacteria in naturally infected fish. The assay is a sensitive, specific, and reproducible diagnostic tool for the simultaneous detection of 4 pathogenic bacteria that cause disease in fish and offers a potentially useful alternative to the conventional culture-based method.

Effects of chronic carbosulfan exposure on liver antioxidant enzyme activities in rainbow trout.

Catalase (CAT), superoxide dismutase (SOD), glutathione S-transferase (GST), and selenium-dependent glutathione peroxidase (Se-GPx) activities in liver of rainbow trout (Oncorhynchus mykiss; 116.88±21.69g) were evaluated after exposing fish to sublethal concentrations (25μg/L) of carbosulfan in flow-through tanks for 60 days. During the experiment activities of CAT, SOD, GST, and Se-GPx and histopathological effects were determined once a week and once at the end of the 21 days of recovery period. All enzymes were affected by carbosulfan when compared to control fish. Fish had intracellular oedema, cell necrosis, pycnotic nucleus, and increase of sinusoidal space in the liver. After 21 days of the recovery period, all enzyme activities had returned to control levels and fish had no histological lesions in liver. Therefore all the changes observed during exposure were reversible. Results indicate that the liver CAT, SOD and GST enzymes are highly sensitive to carbosulfan as their activities altered significantly, suggesting they could be useful in predicting sublethal pesticide toxicity and useful as an indicator for assessment of pesticides in contaminated water.

Effects of Salinity on Yersinia ruckeri Infection of Rainbow Trout and Brown Trout

Abstract Juvenile rainbow trout Oncorhynchus mykiss and brown trout Salmo trutta acclimated to freshwater or salinities of 9.0‰ or less were exposed to Yersinia ruckeri, the bacterial pathogen that causes enteric redmouth disease (ERM). Both species of fish were kept in the same recirculating systems after bacterial exposure. Rainbow trout mortality was significantly (P < 0.05) different in each salinity: 96.5% in freshwater, 89.5% in water of 1.1‰ salinity, 81.3% in 3.0‰ salinity, and 75.0% in 9.0‰ salinity (model SE = 1.0). All deaths occurred between 3 and 12 d after exposure to Y. ruckeri. Only 2.3% of brown trout in all salinities died, and differences among treatments were not significant. For both fish species, Y. ruckeri was isolated from liver, spleen, and trunk kidney of fish dying during this experiment, and lesions of rainbow trout were consistent with ERM. Yersinia ruckeri was not isolated from brown trout surviving for 21 d after bacterial exposure but was isolated from 3 of 24 surviving rai...