This research covers the Master of Science in Fish Biology and genetics, Sylhet Agricultural University, Sylhet-3100, Bangladesh thesis project of T.F.M. Conceptualization, M.A.H., M.M.I. and P.S.; methodology, M.A.H., M.M.I. and P.S.; software, M.A.H. and G.C.; validation, M.A.H., M.M.I. and P.S.; formal analysis, M.A.H. and G.C.; investigation, T.F.M., M.A.H., M.M.I., G.C. and P.S.; resources, M.M.I. and P.S.; data curation, T.F.M., G.C. and A.K.B.; writing—original draft preparation, T.F.M., A.K.B. and G.C.; writing—review and editing, M.A.H., M.M.I. and P.S.; visualization, M.A.H., P.S. and G.C.; supervision, M.A.H., P.S. and M.M.I.; project administration, M.M.I.; funding acquisition, M.M.I. All authors have read and agreed to the published version of the manuscript.

Figure 1. A flow chart illustrating the overall steps of the methodology.

Figure 1. A flow chart illustrating the overall steps of the methodology.

Figure 2. The 96 h Pb(NO3)2 LC50 regression curve for O. niloticus.

Figure 2. The 96 h Pb(NO3)2 LC50 regression curve for O. niloticus.

Figure 3. The mortality rate of O. niloticus in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Figure 3. The mortality rate of O. niloticus in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Figure 4. Hepatosomatic (HSI) indices in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Figure 4. Hepatosomatic (HSI) indices in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Figure 5. Microscopic view of erythrocytes in different treatment groups; (A) Control, (B) T1, (C) T2, (D) T3. (E—erythrocytes, EN—elliptical nuclei; red arrows—shrinking nuclei, black arrows—erythrocytes with rupturing cell membrane, blue arrows—shape deformities); (E,F) Quantitative analysis of erythrocytes in different treatment groups. Different superscripts indicate significant differences at p < 0.05 (CD—cell diameter, ND–nucleus diameter).

Figure 5. Microscopic view of erythrocytes in different treatment groups; (A) Control, (B) T1, (C) T2, (D) T3. (E—erythrocytes, EN—elliptical nuclei; red arrows—shrinking nuclei, black arrows—erythrocytes with rupturing cell membrane, blue arrows—shape deformities); (E,F) Quantitative analysis of erythrocytes in different treatment groups. Different superscripts indicate significant differences at p < 0.05 (CD—cell diameter, ND–nucleus diameter).

Figure 6. Longitudinal microscopic view of gills. (A) Control, (B) T1, (C) T2, (D) T3 (PL—primary lamellae, SL—secondary lamellae, Pc–pillar cells, Ec—epithelial cells, E—erythrocytes, Bc—basal cells, DMC–diffusion of mucous cells, SLD—secondary lamellae damage, EL—epithelial lifting; white circle—acute necrosis, yellow arrows—congestion of basal cells, red arrows—shortening secondary lamellae, black arrows—damage of epithelial layer). Transverse photomicrographs of liver. (E) Control, (F) T1, (G) T2, (H) T3 (Hc—hepatocytes, Nu—nuclei, LD—lipid droplets, LH–liver haemorrhage, NR–nuclear ruptures, DN—degenerated nuclei, MCR—massive cell rupture, V—vacuole; white circle—necrosis, black arrows—cell rupture, yellow arrows—erythrocyte infiltration in blood sinusoids).

Figure 6. Longitudinal microscopic view of gills. (A) Control, (B) T1, (C) T2, (D) T3 (PL—primary lamellae, SL—secondary lamellae, Pc–pillar cells, Ec—epithelial cells, E—erythrocytes, Bc—basal cells, DMC–diffusion of mucous cells, SLD—secondary lamellae damage, EL—epithelial lifting; white circle—acute necrosis, yellow arrows—congestion of basal cells, red arrows—shortening secondary lamellae, black arrows—damage of epithelial layer). Transverse photomicrographs of liver. (E) Control, (F) T1, (G) T2, (H) T3 (Hc—hepatocytes, Nu—nuclei, LD—lipid droplets, LH–liver haemorrhage, NR–nuclear ruptures, DN—degenerated nuclei, MCR—massive cell rupture, V—vacuole; white circle—necrosis, black arrows—cell rupture, yellow arrows—erythrocyte infiltration in blood sinusoids).

Figure 7. Transverse photomicrographs of the intestine. (A) Control, (B) T1, (C) T2, (D) T3. (BB—brush border, AV—absorptive vacuole, LP—lamina propria, L—lumen, EL—extended lumen, IV—increased vacuoles, DAV—disarranged absorptive vacuole; black arrows—tissue rapture, blue arrows—extended serosa, white both side arrows—wider villi); (E) Length and width of intestinal villi in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Figure 7. Transverse photomicrographs of the intestine. (A) Control, (B) T1, (C) T2, (D) T3. (BB—brush border, AV—absorptive vacuole, LP—lamina propria, L—lumen, EL—extended lumen, IV—increased vacuoles, DAV—disarranged absorptive vacuole; black arrows—tissue rapture, blue arrows—extended serosa, white both side arrows—wider villi); (E) Length and width of intestinal villi in different treatment groups. Different superscripts indicate significant differences at p < 0.05.

Table 1. Experimental design with different dosages of Pb(NO3)2 exposure.

Table 1. Experimental design with different dosages of Pb(NO3)2 exposure.

TreatmentConcentration of Pb(NO3)2 (mg L−1)Stocking Density (No./Replicate)ReplicationControl00.00 (00% of LC50)203T105.20 (10% of LC50)203T210.40 (20% of LC50)203T320.80 (40% of LC50)203

Table 2. The 96 h LC50 values for different formulations of Pb in the Oreochromis genus.

Table 2. The 96 h LC50 values for different formulations of Pb in the Oreochromis genus.

SpeciesValue of 96-h LC50
(mg L−1)Formulation of
ToxicantReferencesO. niloticus51.96Pb(NO3)2Current studyO. niloticus40.29[17]O. mossambicus17.33Pb(C2H3O2)2[42]O. mossambicus18.70[43]O. niloticus44.0Pb(NO3)2[44]Oreochromis sp.11.05[18]

Table 3. Physicochemical properties of water in different treatment groups.

Table 3. Physicochemical properties of water in different treatment groups.

ParametersTreatmentDay 0Day 15Day 30Temperature (°C)Control19.93 ± 0.1220.06 ± 0.0420.30 ± 0.21T119.80 ± 0.1019.90 ± 0.0620.20 ± 0.03T219.47 ± 0.0310.87 ± 0.0720.13± 0.08T319.53 ± 0.0719.97 ± 0.1320.20 ± 0.08pHControl8.27 ± 0.068.16 ± 0.058.06 ± 0.03T18.23 ± 0.078.19 ± 0.078.13 ± 0.03T28.23 ± 0.048.2 ± 0.068.17 ± 0.03T38.21 ± 0.078.14 ± 0.068.13 ± 0.07SalinityControl0.07 ± 000.08 ± 0.010.10 ± 00T10.07 ± 0.010.07 ± 0.010.09 ± 0.02T20.06 ± 0.010.08 ± 0.020.10 ± 0.01T30.07 ± 0.030.08 ± 0.010.09 ± 0.01NH3 (mg L−1)Control0.02 ± 0.010.02 ± 0.010.03 ± 0.01T10.01 ± 0.020.03 ± 0.010.03 ± 0.01T20.02 ± 0.010.04 ± 0.010.03 ± 0.01T30.02 ± 0.000.03 ± 0.010.03 ± 0.01DO (mg L−1)Control7.77 ± 0.10 7.06 ± 0.176.96 ± 0.04T18.11 ± 0.07 7.13 ± 0.066.96 ± 0.05T28.05 ± 0.16 6.79 ± 0.026.64 ± 0.09T38.14 ± 0.08 6.57 ± 0.086.73 ± 0.08

Table 4. Growth performance of O. niloticus exposed to Pb(NO3)2 at different concentrations.

Table 4. Growth performance of O. niloticus exposed to Pb(NO3)2 at different concentrations.

ParametersControlT1T2T3Initial length (cm)3.44 ± 0.053.43 ± 0.063.43 ± 0.083.45 ± 0.03Initial weight (g)0.70 ± 0.010.69 ± 0.030.69 ± 0.040.70 ± 0.02Final length (cm)4.40 ± 0.09 c3.82 ± 0.05 b3.70 ± 0.07 ab3.55 ± 0.03 aFinal weight (g)1.47 ± 0.08 c0.78 ± 0.05 b0.55 ± 0.04 a0.52 ± 0.03 aK1.7 ± 0.04 c1.36 ± 0.05 b1.10 ± 0.07 a1.16 ± 0.06 aSGR %2.39 ± 0.20 c0.32 ± 0.26 b−0.77 ± 0.34 a−1.03 ± 0.24 aLength gain %28.35 ± 3.15 c12.08 ± 2.48 b8.23 ± 2.13 ab3.18 ± 0.80 aWeight gain %110.40 ± 11.39 c15.94 ± 8.94 b−13.28 ± 8.72 a−23.60 ± 4.82 a

Table 5. Behavioural abnormalities of fishes among the different treatment units.

Table 5. Behavioural abnormalities of fishes among the different treatment units.

TreatmentDayAbnormalitiesLoss of AppetiteGasping for AirSluggish MovementErratic LocomotionPale GillsSkin Color ChangeControl15------------T1----*------T2***--**T3*******Control30------------T1*********T2**********T3**************

Table 6. Comparative investigation of histopathological alterations from the current experiment in different treatment units.

Table 6. Comparative investigation of histopathological alterations from the current experiment in different treatment units.

OrganAbnormalityControlT1T2T3GillDiffusion of mucous cells---*------ Secondary lamellae damage---***** Epithelial lifting---------** Acute necrosis---******** Congestion of basal cells---*****--- Shortening secondary lamellae------**--- Damage to the epithelial layer------****LiverLiver haemorrhage---******** Nuclear ruptures---******** Degenerated nuclei---------** Massive cell rupture---------*** Vacuole caused by cell rupture---------** Necrosis---******* Cell ruptures---******** Erythrocyte infiltration in blood sinusoids---*******IntestineExtended lumen---------** Increased vacuoles---------*** Disarranged absorptive vacuoles------**** Tissue rapture---*** Extended serosa---------* Wider villi---*****