Clinicohematological, Mutagenic, and Oxidative Stress Induced by Pendimethalin in Freshwater Fish Bighead Carp (Hypophthalmichthys nobilis)

Abstract

Currently, aquatic and terrestrial ecosystems are continuously and chronically polluted by cocktails of countless chemical compounds. The susceptibility to infections is tremendously increasing in a variety of organisms due to exposure to environmental pollutants. Pendimethalin, an herbicide, is continuously used in agriculture to remove unwanted broadleaf weeds across the globe. Therefore, this study investigates the mechanisms of toxicity of pendimethalin in freshwater fish bighead carp upon exposure to low and environmentally relevant concentrations. For this purpose, 48 fish without any clinical abnormalities were kept in a glass aquarium in different experimental groups (T0, T1, T2, and T3). These groups were treated with pendimethalin at 0.00, 0.25, 0.50, and 0.75 mg/L, respectively. Four fish were randomly picked from each experimental group and killed at 72, 96, and 120 hours of the trial to study hematobiochemical parameters and visceral tissues including the brain, liver, heart, gills, and kidneys for histopathology. Herbicide-treated fish indicated various physical and behavioral abnormalities including hypersecretion of mucus, erratic swimming, operculum movement, air gulping, tremors of fins, loss of equilibrium, and increased surface breathing. Histopathologically, gills tissues of treated fish indicated atrophied lamellae, uplifting of secondary lamellae, necrosis of primary and secondary lamellar epithelial cells, telogenesis, congestion, and lamellar fusion. Histopathological examination of liver tissues of treated fish showed mild to moderate congestion, necrosis of hepatocytes, and atrophy of hepatocytes while kidneys revealed degeneration of renal tubules, glomerular atrophy, ceroid, and necrosis of renal tubules. The erythrocyte counts, monocyte and lymphocyte counts, and hemoglobin values were significantly (P < 0.05) reduced in pendimethalin-treated fish. Results on serum biochemistry showed that the biomarkers of kidneys, heart, and liver were significantly higher in fish of treated groups. In addition, values of different biochemical reactions like reactive oxygen species (ROS), thiobarbituric acid reactive species (TBARS), total proteins, and quantity of different antioxidant enzymes including reduced glutathione (GSH), catalase, and superoxide dismutase (SOD) were significantly different when compared to untreated fish. Moreover, the percentile of different nuclear abnormalities in red blood cells and frequency of DNA damage increased significantly in treated fish. It can be concluded from the findings that pendimethalin causes its toxic effects via disruption of physiological and hematobiochemical reactions of fish.

Figure 1

Photomicrograph of Hypophthalmichthys nobilis (bighead carp) treated with pendimethalin (0.75 mg/L) for 120 hours (a) blood smear stained with Giemsa-Wright stain (1000x) showing micronucleus inside the erythrocytes (arrowheads), abnormally shaped erythrocytes (A), pear-shaped erythrocytes (P), and elongated (E) erythrocytes (upper photo) and (b) DNA damage fluorescing of nuclear material around the cells (lower photo).

Figure 2

Trend in DNA damage in (a) lymphocytes/hepatocytes and (b) kidney cells and gills in Hypophthalmichthys nobilis (bighead carp) treated with pendimethalin with different doses (T0: control; T1: 0.25 mg/L; T2: 0.50 mg/L; T3: 0.75 mg/L) at different time intervals. Bars (mean ± SE) bearing asterisk differ significantly (P ≤ 0.05) in comparison with other treatments.

Figure 3

Photomicrograph of the brain of Hypophthalmichthys nobilis (bighead carp) treated with pendimethalin (0.75 mg/L) for 120 hours showing (a) necrosis of neuron, atrophy of neuron, degeneration of cytoplasm of neuron, and microgliosis; (b) necrosis of neuron, atrophy of cytoplasm, and microgliosis; and (c) cytoplasmic atrophy, degeneration of neuron, and necrosis. Hematoxylin and eosin stain: 400x.

Figure 4

Photomicrograph of gills of Hypophthalmichthys nobilis (bighead carp) treated with pendimethalin (0.75 mg/L) for 120 hours showing (a) disruption of cartilaginous core (arrow), uplifting of secondary lamellae, and necrosis of lamellar epithelial cells and (b) disorganization of cartilaginous core and vacuolation (arrowheads). Hematoxylin and eosin stain: 400x.

Figure 5

Photomicrograph of kidneys of Hypophthalmichthys nobilis (bighead carp) treated with pendimethalin (0.75 mg/L) for 120 hours (a–c) showing severe necrosis and detachment of tubular cells from the basement membrane (arrowheads), infiltration of inflammatory cells in between the renal tubules (arrows), and widening of urinary spaces. Hematoxylin and eosin stain: 400x.