International Journal of Aquaculture, 2018, Vol.8, No.14, 104-111
106
1 Materials and Methods
1.1 Experimental site
Experiments were conducted at the Wet Laboratory and Fish Conservation Laboratory under the Department of
Fisheries Management of the Faculty of Fisheries, Bangladesh Agricultural University (BAU), Mymensingh.
Active and healthy specimens of striped gourami fish (
Trichogaster fasciata
) were randomly collected and
stocked in the cisterns of Mini hatchery and Breeding complex located at the southern side of the Fisheries
Faculty building. The size of each cistern was 250 cm ×195 cm ×70 cm. Aeration was maintained and the fishes
were kept under natural photoperiod. Commercial fish feed were fed twice daily to satiation.
1.2 Experimental design for gills histology
After acclimatization in the cisterns the fishes were exposed to different concentrations of chlorpyrifos 20 EC
for different times to find the LC
50
value and to identify the histopathological changes of gills. Using
Probit-Analysis LC
50
of chlorpyrifos for striped gourami was determined. There are five sub-lethal doses, such as
0, 15, 50, 150 and 500 µg/L of chlorpyrifos which were denoted as T
c
(control), T
15
, T
50
, T
150
, and T
500
respectively. There are five treatments and 3 replications in each treatment. Each of the concentrations and control
group was maintained in triplicates. To conduct this experiment, 15 PVC tanks (86 cm diameter, 81 cm depth)
were used and cleaned with disinfectant, washed thoroughly with ground water. After that each of the tanks was
filled with 300 L of dechlorinated ground water. Then 10 female (length 7.97 ±0.72 cm and weight 9.76 ±2.31 g)
fishes were kept in each of the 15 tanks which were identified by their silvery body color and swollen abdomen.
Sub-lethal doses were applied according to LC
50
of the chlorpyrifos which was estimated for adult striped
gourami. After 21 days of conditioning, the sub-lethal concentrations were exposed to 15 tanks for 15, 30, 45, 60
and 75 days. About 90% of tank water was exchanged every alternate day and fresh chlorpyrifos was used. Fishes
were feed twice daily and excess food and excretion were removed through siphoning every day. After 15 days
interval, female fishes were sampled regularly. Total length and body weight of each fish was determined using a
specialized scale.
1.3 Histology of gills
The collected fish were taken on a tray and operculum was cut carefully by scissor and gills were exposed. Gills
were removed with the help of scissor and placed on a petridish. Then the samples were preserved in 10% neutral
buffered formalin for further analysis. The fixed gills were passed through graded alcohol series to dehydrate them.
The dehydrated gills samples were taken for clearing with 100% benzene for 2 times with an interval of 1 hr at each
step. The samples were embedded into paraffin. Sectioning was done using microtome. The gill sections were then
stained for 2 times with haematoxylene and eosin stains. Finally the gills sections were observed under microscope.
1.4 Microscopic observations
The slides were observed under electric microscope (Olympus) which was connected to computer with a viewer
(Magnus viewer). The viewer was also equipped with a camera. By the help of this mechanism numerous
photographs were snapped at different magnifications.
2 Results
Histopathological results indicated that gill was the primary target tissue affected by chlorpyrifos. The gills are
key organs involved in nutrient uptake, ingestion and respiration. The gill tissue of
Trichogaster fasciata
in
general consists of well structured primary and secondary gill lamellae. This is evident in gill tissues not exposed
to chloropyrifos (control). Continuous exposure to the chlorpyrifos caused architectural distortion of the gill
tissues of the exposed fish as shown in figures (sub lethal concentrations). No histological changes were observed
in the gill of the control fish (0 μg/L). The most common changes occurred in a range of 50 μg/L-150 μg/L
concentrations of chlorpyrifos were hypertrophy, necrosis, missing of gill lamellae, hemorrhage, vacuum,
pyknotic cells and splitted of gill lamellae.
The gills exhibited chloropyrifos induced changes on the 15
th
day exposure (Figure 3). Almost normal structure
appeared when exposed to 15 μg/L. But significant structural change like hypertrophy was observed at 50 μg/L
