1. science
2. ecology
3. pollution

Journal of Academia and Industrial Research (JAIR)
Volume 3, Issue 11 April 2015
562
ISSN: 2278-5213
RESEARCH ARTICLE
Acute Toxic Effects of Hevea brasiliensis on the Gills of Hatchery Reared
Oreochromis niloticus Fingerlings
1
George Ubong1*, Ini-Ibehe N. Etim2, M.P. Ekanim2 and Akpan, Monica K.2
Dept. of Fisheries and Aquaculture, Institute of Oceanography, University of Calabar, Cross River State, Nigeria
2
Dept. of Marine Biology, Akwa-Ibom State University, P.M.B. 1167, Uyo
[email protected]*; +2348032625310
______________________________________________________________________________________________
Abstract
The effects of water soluble fraction (WSF) of Hevea brasiliensis on the gills of Oreochromis niloticus
fingerlings had severe impacts on the test organisms. Six concentrations ranging from 0, 8, 16, 24, 32 and
40 mg/L were prepared from the WSF of the latex for histopathological examination. There was no
observable change in the gills of the test organism in the control group. The test organism in the 8, 16, 24,
32 and 40 mg/L concentration exhibited various reactions which included irregular movement, attempt at
jumping out, color changes, exhaust swimming motions and changes in opercula rate. In the 8 mg/L
concentration, the gill showed loss of cells integrity, in the 16 mg/L concentration it was observed to show
loss of cell integrity and disintegration, in the 24 mg/L of toxicant, cell disintegration was observed in the gills,
in the 32 mg/L of toxicant loss of gill secondary lamellae was observed and in the 40 mg/L concentration
pronounced lamellae erosion and hyperplasia was observed in gill ray. From the findings, it is observed that
latex obtained from Hevea brasiliensis is toxic to aquatic life. Therefore, effective management strategies
should be put in place to ensure safety compliance.
Keywords: Hevea brasiliensis, Oreochromis niloticus, histopathology, toxicant, management strategies.
Introduction
Despite the numerous benefits that are rendered to the
modernization of this world by natural rubber, the
consequence of natural rubber processing has yet to
provide a serious problem due to its highly polluted
effluents. The rapid growth of this industry generates
large quantities of effluents coming from its processing
operations which is really a big problem because its
wastewater contains ammonia and high biological
oxygen demanding substances. These substances can
deplete dissolved oxygen and create anaerobic
conditions in water bodies. Without proper treatment,
discharge of wastewater from rubber processing industry
to the environment may cause serious and long lasting
consequences (ATPC, 2004). Pollution has become a
general term for the common man because of its regular
occurrence. Environmental pollution has been woven into
the fabric of our modern life. Very often, the world is
shocked by reports of pollution disasters and then man
becomes active and conscious about the harmful effects
of pollution and thinks about the ways and means to
keep the environment clean. Studies have shown that
structural changes in gills are induced by toxicant
(Nowak, 1992; Tamse et al., 1995). From results
obtained from previous studies, the damage to gills as a
result of toxicant effects strongly suggests that the gill is
a common mode of action by which hyperventilation is
induced because most of these histological changes
increase the diffusion distance for oxygen from the water
to the blood, which again could explain the separation of
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the respiratory epithelium. Acute exposure to some toxic
chemicals can result in severe destruction of the gill
lamellae within a few hours and further affects the
rate of oxygen consumption (Health, 1995). The study of
Environmental toxicology is thus, concerned with how
Environmental toxicants, through their interaction with
humans, animals and plants, influence the health and
welfare of these organisms. Ecotoxicological studies are
very vital because it provide information on the
concentration of a particular toxicant (i.e. at what level of
concentration it is lethal or sub-lethal) and also helps in
understanding the adverse effects associated with a
particular toxicant. According to Azevedo-Santos et al.
(2011), Oreochromis niloticus is a species of tilapia, a
cichlid fish native to Africa and as far west as Gambia.
It is also native to Israel and numerous introduced
populations exist outside its natural range (e.g. Brazil).
The choice of Oreochromis niloticus as a biological
indicator in this study correlates with the fact that it is an
important aquaculture species in many part of the World,
including Nigeria. The present investigation was
conducted to provide information on the histological
changes in the gills of Oreochromis niloticus under
exposure to Hevea brasiliensis.
Materials and methods
Test organism: Hevea brasiliensis was collected from the
Rubber research institute, Calabar, Nigeria located within
Calabar (08o22’10.583’E and 05o8’2.85’’N) (George
et al., 2014) (Fig. 1).
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Ubong et al., 2015
Journal of Academia and Industrial Research (JAIR)
Volume 3, Issue 11 April 2015
563
Fig. 1. Map of the study area.
Definitive test: The final concentration chosen for the
WSF of Hevea brasiliensis latex for the toxicity test on
Oreochromis niloticus after the preliminary tests ranges
from 0, 8, 16, 24, 32 and 40 mg/L. The experiment lasted
for 96 h. The fish were not fed in order to reduce waste
production. Distress behavior were closely monitored
and recorded from the onset of the experiment for 6, 12,
24, 48, 72 and 96 h respectively. Prior to stocking,
initial water parameter were monitored and also
daily water parameter such as dissolved oxygen,
temperature, pH, nitrite and ammonia were monitored
using mercury-in-glass thermometer, Lurton Do and pH
meters and nutrafin test kits for measuring the
concentration of ammonia and nitrite. The battery
operated meters were calibrated according to
manufacturer’s instructions before being used for
measurement (Boyd, 1989, 1990).
About 120 healthy Oreochromis niloticus fingerlings in
the range of 2-4.5 cm in size procured from the
University of Calabar Fish Farm, Cross River State
located within the University of Calabar at latitude 0405,
020’N and longitude 008020’ 450’E, respectively (Asuquo
and Bassey, 1999; Akpan et al., 2002) were used for the
studies and was transported to the Research Laboratory
of the Institute of Oceanography (IOC), University of
Calabar for acclimatization. Oreochromis niloticus
fingerlings were acclimatize to laboratory conditions for
24 h in the glass tank and aerated with air stone
connected to electrically powered aquarium pumps for
the test organisms to get acquainted with the
environment.
Preparation of toxicant solution: Water soluble fraction
(WSF) of Hevea brasiliensis latex was obtained by
vigorously shaking rubber extract with filtered habitat
water in a separatory funnel. The system was allowed to
stand for 6 h for complete phase separation, after which
the lower aqueous layer containing the WSF was
decanted for the toxicity test.
Stocking of specimens: Twelve glass tanks measuring
25 x 10 x 15 cm was used for the toxicity test. The glass
tank was filled with 2 L of dechlorinated water each.
Ten Oreochromis niloticus fingerlings were gently picked
using a hand net in order to avoid stress into each of the
glass tank
Range finding test: The concentration chosen for the
range finding test of WSF of Hevea brasiliensis latex on
Oreochromis niloticus ranges from 0, 10, 20, 30, 40 and
50 mg/L. Ten fishes were randomly picked and
introduced into each of the reconstituted latex and each
concentration was set in duplicate with control containing
dechlorinated water without the presence of WSF of
Hevea brasiliensis latex.
©Youth Education and Research Trust (YERT)
Histopathology: Tissues (gills) isolated from the test
animal were fixed in formal-saline for 48 h. The fixed
tissue (gills) were processed manually through graded
ethanol, cleared in xylene, impregnated and embedded
in paraffin wax. These sections were cut with a rotary
microtome, stained by haematoxylin and eosin
technique, studied microscopically for pathological
changes.
Statistical analysis: Statistical analysis was done by
comparing the difference between control and Hevea
brasiliensis treated groups by students t-test using
(SPSS Inc; Chicago, USA). The results were considered
significant (P<0.05).
Results
Various reactions were exhibited by the test organism
which included, weaken swimming motions, changes in
opercula rate, erratic movement, vertical swimming
position, attempt at jumping out and color changes.
The mucous secretion increased and accumulated on
the gills. Hemorrhagic patches appeared on the ventral
surface of the fish with general discoloration and anoxia.
Gill epithelium of fish from the control group was similar
to that of other teleostean fishes. However, tilapia
exposed to WSF of Hevea brasiliensis showed several
alterations in the gill (Table 1).
Table 1. Histological changes in gills
of O. niloticus fingerlings.
Conc.
(mg/L)
0
8
16
24
32
40
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Histological changes
Normal distribution of gill lamellae
Loss of cells integrity in the gills
Loss of cells integrity and disintegration of the gills
Cells disintegration
Loss of gills secondary lamellae
Lamellae erosion and hyperplasia
Ubong et al., 2015
Journal of Academia and Industrial Research (JAIR)
Volume 3, Issue 11 April 2015
564
Plate 1. Normal distribution of gill lamellae at 0 mg/L of toxicant (X100).
Plate 5. Loss of gill secondary lamellae at 32 mg/L of toxicant (X100).
Plate 6. Lamellae erosion and hyperplasia of gill at
40 mg/L of toxicant (X100).
Plate 2. Loss of cell integrity in the gills at 8 m/L of toxicant (X100).
Plate 3. Loss of cell integrity and disintegration in the gills
at 16 mg/L of toxicant (X100).
Plate 4. Cell disintegration in the gills at 24 mg/L of toxicant (X100).
No recognizable changes were observed in the gills of
control fish (Plate 1). In contrast with the control ones,
gills in fish exposed to Hevea brasiliensis was found to
induce several histological alterations in the gills from
slight to deep alterations, which were found to be
concentration-dependent. After exposure to 8 mg/L of
Hevea brasiliensis, the sections of fish gills showed loss
of cells integrity (Plate 2). Exposure to 16 mg/L of Hevea
brasiliensis showed loss of cell integrity and
disintegration (Plate 3). In Plate 4, the gills of the fish are
shown to have undergone cells disintegration at
exposure to 24 mg/L of the toxicant, while Plate 5 shows
loss of gills secondary lamellae at exposure to 32 mg/L
of toxicant and at Plate 6, the gills of the fish was
observed to undergone moderate to extensive lamellae
erosion and hyperplasia.
Discussion
In aquatic ecosystem, industrial effluents are considered
as one of the most important pollutants, since they
find their way into the ecosystem through deliberate
discharge or surface run-off. Once discharged into water
bodies, they can either be adsorbed on sediment
particles or accumulated in aquatic organisms. Fish may
absorb dissolved elements from surrounding water and
food, which may accumulated in various tissues in
significant amounts and are electing toxicological effects
at critical targets. This bioaccumulation may lead to high
mortality rate or cause many histological alterations in
the survived fish organs.
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Ubong et al., 2015
Journal of Academia and Industrial Research (JAIR)
Volume 3, Issue 11 April 2015
565
This led us to undertake the study of Hevea brasiliensis
toxicity to Nile Tilapia basing on histological studies of
gills after exposure to water soluble fraction of these
compounds. Histological study of the gills of Nile tilapia
(Oreochromis niloticus) showed a typical structural
organization of the lamellae in the control group.
However, fish exposed to water soluble fraction of
H. brasiliensis latex showed several histological
alterations of the gills of the exposed organism. In the
8 mg/L concentration of the toxicant, it was observed to
cause loss of cell integrity in Oreochromis niloticus.
In the 16 mg/L of the toxicant, the effects of
H. brasiliensis were observed to cause loss of cells
integrity and disintegration in Oreochromis niloticus
fingerlings. In the 24 mg/L concentration of the toxicant,
the gill lamellae of Oreochromis niloticus were observed
to undergo cell disintegration. Loss of gills secondary
lamellae was observed in 32 mg/L concentration of the
toxicant. Lamellae erosion and hyperplasia condition was
observed in the gills of O. niloticus at 40 mg/L.
Gill lamellae cell disintegration has been reported by
Diana et al. (2007) in Carassius auratus gibelio when
investigating biochemical and histological effects of
deltamethrim on the species with different effects such
as lamellae cells hypertrophy and nuclear pycnosis
in the basal cells. Hypertrophic, necrotic, atrophy and
dystrophy of secondary lamellae have also been
reported in Clarias gariepinus juveniles exposed to
refined petroleum oil and kerosene under laboratory
conditions. These histological alterations have been
observed by several authors in fish submitted to several
toxicants (Karan et al., 1998; De Boeck et al., 2001).
The general morphological changes in the gills recorded
in this study have been reported in Astyanax sp. After
96 h exposure to water soluble fraction of crude oil
(Akaisha et al., 2004). Gabriel et al. (2007) also reported
similar changes in Clarias gariepinus exposed to
petroleum oil and kerosene, reported in Oreochromis
niloticus by Wannee et al. (2002), Ayoola (2011) and
Ekpo and Chude (2011). Generally, fish gills are the
prime target organ of all pollutants due to their sensitive
and net-like structure. Both dissolved oxygen and
colloidal materials tend to adhere to fish gills resulting in
superficial rupture and disease condition to the gills.
Gill morphology and configuration are important
biomarkers, providing a basis for the detection of
pollutants. Gills are considered as the most vulnerable
organ to the changes in the aquatic environment (Bernet
et al., 1999), because they are in direct contact with the
surrounding water that may contain Hevea brasiliensis
and consequently, they are the first entrance door for
these and other environmental pollutants (Khan and
Kiceniuk, 2003). Gill is an important environmental
indicator providing a quick means of detection of the
effects of pollutants. Gills of the fish exposed to the
toxicants showed several histological alterations as a
result of the toxicant, this was also reported by Camargo
and Martinez (2007).
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Gills are very sensitive and respond extremely fast to
water pollution caused by Hevea brasiliensis (George
et al., 2014). The basic consideration of the use of gill of
Oreochromis niloticus as environmental indicator of
pollution is that living organisms provides the best
reflection of the actual state of ecosystem and changes
therein. Results obtained can provide a reasonable basis
for comparing the effects of such pollutants on higher
animals and human beings.
Conclusion
In the present study, the effects of the toxicant on the
gills of the test organism were concentration-dependent.
The higher the concentration, higher the manifestation
effects of the toxicant on the gills of the test organism.
Similar report was presented by George et al. (2014)
when investigating toxicological impact of Hevea
brasiliensis on the gills of fingerlings of African catfish
Clarias gariepinus. Our findings uphold the concept of
Mallat (1985) that toxicant induced alteration in gill
histology is largely non-specific, because similar types of
lesion occur under a wide range of toxicant-exposure
conditions.
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