The most reliable and surer way of fingerling production is induced breeding.
According to Viveen et al. (1985) fish fingerling production involves
a series of breeding and feeding activities which result in the mass production
of fish seeds under controlled conditions and environment, usually in
a hatchery. One of these activities is induced spawning which involves
the use of hormones (synthetic or non-synthetic). The hormone is meant
to hasten the ripening of the eggs within few hours. The various hormones
in use include pituitary extract or hypophysis from similar or different
fish, deoxycorticosterone acetate (DOCA), Ovaprim and human chronic gonadotropin.
In Nigeria these synthetic hormones are not readily available and are
very expensive (Adebayo and Fagbenro, 2004).
Hypophysation, the use of pituitary extracts is presently the most popular.
However, they are uneconomical and wasteful because adult fish are sacrificed
(Salami et al., 1994). Apart from the fish pituitary which is the
popular choice, the frog pituitary can also be used (Fagbenro et al.,
1992). In Nigeria, frogs often constitute nuisance in ponds and around
wetlands particularly during the rainy season. They could therefore be
put to a reasonable use by making use of their pituitary extracts and
storing them against the dry season when their population drastically
reduces. This is possible, if only adequate information abound concerning
the viability of the pituitary extracts after being stored. This will
prevent farmers from spending so much on ovaprim and fish which are usually
sacrificed (killed) when their pituitary extract is needed.
Researches have been carried on the potency of pituitary extract of frog
(Moustafa et al., 1984; Fagbenro et al., 1992) and the various
researchers have proved that the pituitary extract of frog is viable for
induced breeding of fish. However, the effect of storage on the potency
of the extract of frogs` pituitary has been hitherto unexplored.
The objective of this study therefore, was to examine the effect of varying
storage period on the efficacy of frog pituitary with the aim of determining
the optimum period required for effectively enhancing induction of ovulation,
maturation and artificial propagation of Clarias gariepinus.
MATERIALS AND METHODS
Hormone Collections and Preparation
The hormone use in this study was acetone dried pituitary extract
from the African bullfrog, Rana adspersa. Mature donor African
bullfrogs weighing > 120 g were collected over a period of 4 months
(April to August 2006) within the Fish Farm of Federal University of Technology,
Akure (FUTA), Nigeria.
The frog was killed by a blow on the head using a wooden mallet (Fagbenro
et al., 1992), headed and the skull was opened. The pituitary is
located in the skull. The palate of the mouth was opened with a pair of
pincers and from there the pituitary was collected.
After collection, the pituitary extract was stored dry following the
procedures described by Viveen et al. (1985). The pituitary was
put in a vial filled with some acetone (1 mL acetone per pituitary) and
was refreshed after 10 min. The acetone was again renewed after 8 h. It
was then completely drained after 24 h and the pituitary was dried in
shade by evaporation and stored in a sealed vial placed in a desiccators.
Each of the vials was labeled in weeks.
Brooder Selection and Maintenance
Mature male and female C. gariepinus broodfish were procured
from a reputable farm in Akure (Nigeria) and stocked in earthen ponds
on Federal University of Technology, Akure fish farm. Sexually matured
adults (average weight 500 g) were selected on the basis of their external
sexual characteristics. Mature males had a prominent vascular red pointed
genital papilla, while the female had a soft, highly distended abdomen
with a red swelling around the cloaca.
Female brooders used for induced spawning trials were further selected
using ovarian catheterization (in vivo determination of the stage
of intra-vision oocytes). Oocyte diameter was measured with a micrometer
using a binocular microscope (x25) and brooders that produced oocytes
diameter of at least 1.1 mm were selected, tagged and later transferred
to outdoor rectangular cement cisterns (5x4.5x1.5 m)
Preparation of Hormones
Acetone dried pituitary extracts for each month were crushed into
a fine powder, in a porcelain-mortar after which 2 mL of 0.9% NaCl solution
was added and thoroughly mixed. The females were injected intramuscularly
above the lateral line away from the head in the direction of the tail
at angle 45 °C. The males were not given any doses.
Induction of Ovulation, Fertilization and Incubation
Fifteen spawning trials were carried out i.e., 3 trials per treatment
(with the storage period serving as the treatment) over the 16 week period.
Fish were placed in individual covered container. The female fish so
injected were removed after 12 h and eggs were stripped into a dry plastic
bowl. Males were dissected and their testis removed and the milt used
to fertilize the eggs following the procedure described by Viveen et
al. (1985). A portion of the eggs from each fish for each month and
the control was collected in a bowl and kept in the FUTA Hatchery to determine
the percentage fertilization, hatchability and survival. The larvae were
fed shell-free artemia for one week after yolk absorption.
All percentage data were arc sine transformed prior to analysis. Data
obtained were pooled for each treatment and compared by one-way analysis
of variance (ANOVA) test to determine significant differences (p = 0.05)
and Turkey`s post hoc test was used to determine differences among treatment
RESULTS AND DISCUSSION
Generally the value obtained for the ovulation and spawning responses
in the hormone treatment over the various storage periods were significantly
different (p<0.05). It was observed that the storage period had effect
on the ovulation of C. gariepinus (Table 1).
The optimum storage period was 4 weeks. The relative fecundity, fertilization,
hatchability and survival rate were highest for fresh frog pituitary extract
as shown in Table 1. This may be due to the fact that
all the chemical constituents of the hormone have not in any way been
tampered with or altered.
The results of this study indicated that the yield of R. adspersa
pituitary hormone extract stored for 4 weeks was significantly different
from that gotten from hormones stored for over 8 to 16 weeks period. It
showed that the frog pituitary extract is still potent, viable and effective
when stored for not more than 4 weeks.
The mean weight of eggs spawned (per kilogram fish) using hormones stored
for 12 and 16 weeks were low in relation to the weight of the fish used.
Fecundity was also low relative to the weight of fish used. Fertilization
rate was high in the control hormones treatment and at 4 week - storage
period (>60%). These values were however very low in fish administered
with the hormones stored for 8 weeks while it was negligible for fish
treated with hormones stored for the 12 and 16 weeks period.
Fresh pituitary treatment shows fertilization of 92%. This spawning response
for fresh pituitary treatment is comparable with that reported by Fagbenro
et al. (1992). They reported that C. gariepinus were successfully
spawned using frog pituitary hormone with an average fertilization of
98%. The result obtained for fish given the fresh pituitary extract for
ovulation induction were significantly different from the rest generally.
The spawning response of C. gariepinus to pituitary extract of
R. adspersa stored for 4 week is similar to those reported for
other hormones, such as human chorionic gonadotropin for Heterobranchus
bidorsalis (Legendre, 1986) and C. gariepinus (Mollah
and Tan, 1983). Hatching time and the latency period for all the treatments
were however not significantly different (p>0.05) from one another.
The injection of FPE stored for 0-4 weeks elicited high ovulation response
with latency period of 12 h at 26 ± 1 °C. This is in agreement
with latency period reported by Britz and Hecht (1998) for C. gariepinus
at 25 °C using C. gariepinus pituitary extract. The
latency period for this study is much shorter than 15-18 h reported by
Adebayo and Fagbenro (2004) for Heterobranchus bidorsalis at 27
± 1 °C using carp pituitary extract and C. gariepinus
pituitary extract. The hatching time ranged between 24 to 26 h under the
prevailing temperature of 25-27 °C. This is in line with Viveen et
al. (1985) who reported hatching time of 23-27 h for the temperature
||Induced ovulation and Spawning of C. gariepinus using
frog pituitary extract stored for over a period of 16 weeks
|Values are means ± SD from three replicates.
Means in each row with different superscripts are significantly different
The results of this study suggest that time factor play a major role
in the efficacy of frog pituitary hormone. This study is important as
it showed that although there is still a significant different in the
overall result obtained over the various storage period, yet hormone stored
for 4 weeks still gave a fairly yield in terms of fertilization, hatchability
and survival. Farmers can therefore afford to store their pituitary for
a maximum of 4 weeks for induced spawning of C. gariepinus.