1. 1
EBONYI STATE UNIVERSITY
SCHOOL OF POST GRAGUATE STUDIES
APPLICATION FOR APPROVAL OF TITLE OF THESIS
NAME OF STUDENT EZIKE Christopher Onyemaechi
REGISTRATION NUMBER EBSUPG/Ph.D/07/01901
DEPARTMENT Fisheries and Aquaculture
FACULTY Agriculture and Natural Resources Management
DEGREE IN VIEW Ph.D
EXPECTED YEAR OF GRADUATION 2014
PROPOSED TITLE OF THESIS Effects of Water Soluble Fraction of Crude Oil on
the Haematology, Histopathology and
Carbohydrate Reserves of Clarias gariepinus
(Burchell, 1822) Juveniles
SYNOPSIS
INTRODUCTION
Pollution from crude oil is common occurrence world over, particularly in nations whose
economies are largely dependent on the oil industry. This is the case with Nigeria where it has
remained the most important source of revenue generation for the past four decades. Oil
exploration and exploitation at the onshore and offshore are carried out at the Niger Delta areas,
producing more than 90% of crude oil in Nigeria and thus hosting most of the terminals of oil
(Wegwu and Omoedu, 2010). Spill incidences of various scales have been reported at the
swampy and coastal areas of the Niger Delta (RSEPB, 1992). Spillage occurrence are regular
features in the oil producing communities of the region due to leakages from numerous pipelines,
criss-crossing the sensitive ecological terrains and aquatic environment of the oil communities,
hose failure, sabotage, vandalization, burst arising from rupture of flow lines, accidents from
loading and shipping activities, drilling operations, blow up of oil rigs among other factors that
cause spillage in these areas (Omoregie, 2002). Measurement of acute mortality effects and
subacute physiological effects of the water soluble fraction (WSF) of Bonny Light crude oil
(BLCO) on the haematology, histopathology and changes in the carbohydrate food reserves of
Clarias gariepinus are important biornarkers for assessing toxicant effects on exposed fish. The
specific objectives of this study are to: (i) determine the 96hr LC50 and safe concentration of WSF
of Bonny Light crude oil on Clarias gariepinus juveniles (ii) evaluate the effects of water soluble
fraction of Bonny light crude oil on the haematology of juveniles of Clarias gariepinus (iii)
determine the effects of water soluble fraction of the Bonny light crude oil on the histopathology
of juveniles of Clarias gariepinus (iv) evaluate the effects of water soluble fraction of Bonny light
crude oil on the carbohydrate reserves of Clarias gariepinus.
MATERIALS AND METHODS
Seven hundred and fifty (750) juveniles of Clarias gariepinus (initial mean weight 8.61±0.02g
and length of 11.37±0.076cm) from same brood stock and age were collected from Luis Farm,
Warri, Nigeria and transported in plastic containers to the Departmental (Wet) Laboratory.
Thereafter the fish were acclimated to laboratory conditions in 50litres capacity plastic vats
for 14 days and fed at 3% body weight, twice daily using 38% crude protein experimental diet.
Preparation of water soluble fraction of crude oil was carried out with galincarp stirrer rotated for
20h by a magnetic stirring rod. The experimental scope consisted of: 24hours range finding test
(RFT) to determine acute concentration ranges, Definitive test (DT) - 96hours acute exposure at
dose levels of 0.00, 50, 100,150, 200 and 250ml/l of WSF of BLCO exposed to10 juveniles per
treatment (replicated thrice) in 50liters capacity vats to determine 96h LC50, safe level, and
behavioral changes and 10weeks subacute exposure at 0.00, 1.25, 2.5, 5, 10 and 20ml/l of WSF
of BLCO exposed to 20 juveniles per treatment and replicated thrice to determine effects on
haematology, histopathology and carbohydrate reserves of exposed fish. Concentration with

2. 2
0.00ml/l of WSF of BLCO served as control. Water quality parameters were monitored daily during
the DT and at 2weeks intervals during subacute exposure. Fish was inspected respectively at hourly
and daily basis for mortality during RFT and DT. From start to the 10 weeks subacute exposition, 2
fish from each treatment were sampled every 2 weeks and blood and dissected vital organs (gills,
liver, kidney and muscle) were respectively taken for analysis of haematology, histopathology and
carbohydrate reserves. Determination of blood parameters were carried out in a standard
haemacytometer (model JSQA). Histological examination of damaged tissues were carried out
with automatic tissue processor (model Leica AP) and same were sectioned with rotatory microtome
(model Leica RM) and stained'with eosin and hematoxylin while reserves of carbohydrate were
estimated using the Anthrone method.
RESULTS
Results indicate as follows (i) exposure of Clarias gariepinus juveniles to acute concentrations of
WSF of BLCO above threshold level of 83.18ml/l did not affect water quality parameters but
produced signs of toxicosis and death of 50% of exposed fish at 96h LC50 of 109.95ml/l and safe
level of 1.1 ml/I (ii) lower subacute levels of WSF of BLCO did not cause mortality but resulted as low
as 1.25ml/l to reduced number of white blood corpuscles (WBC), which led to increased neutrophil
levei, however red blood corpuscles (RBC), packed cell volume (PCV) and mean blood
haemoglobin suffered reduction from 2.5ml/l while mean number of lymphocytes reduced
significantly from 10ml/l of toxicant. (iii) Histopathology examination revealed reversible cell
responses from 1.25 to 5ml/I WSF of BLCO but irreversible tissue damages began from 10ml/l
exposure and resulted to tissue atrophy in kidney and muscle, massive vessel damages in liver
coupled with cirrhosis of the same and talengiectasia of secondary lamellae of gills. Necrosis of
muscles and liver hepatocytes, erosion of secondary lamellae of gills and apoptosis in kidney tissues
were observed in fish exposed to 20ml/l of WSF of BLCO. (iv) lower levels of toxicant reduced
carbohydrate food reserves in which liver and muscle glycogen were decreased significantly while
plasma glucose reduced significantly at 1.25ml/l of WSF of BLCO and resulted to hypoglycaemia but
the same increased significantly from 5mI/I and resulted to hyperglycaemic condition in exposed fish.
---------------------------------------------- --------------- ---------------------- -----------------
EZIKE, Christopher Onyemaechi Date Prof. Ugwu, L.L.C Date
Student Supervisor
---------------------------------------------- --------------- ---------------------- -----------------
Dr. Ude, E. F. Date Dr. Umeh, G.N. Date
H.O.D., Fisheries and Aquaculture Fac. Rep., SPGS

3. 3