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Thesis Issued 2026-09-08 EN

Production of yy supermale of nile tilapia (Oreochromis niloticus) and cryopreservation of sperm

Author: Muhammad Zahirul Islam

Abstract

A series of experiments were conducted to identify and propagate YY supermales of tilapia from YY paternal stock collected from Central Luzon State University of Philippines. At first, 10 fish were bred individually with normal females (XX) and nine fish were identified as putative YY males which produced 86.7% to 99.0% male progeny. Four putative YY males were bred separately with normal females (XX) and the produced XY fry were feminized with 200mg of ẞ-estradiol per kg of feed which produced 93.4% to 98.0% females. These ẞ-estradiol treated females were reared upto maturity and 24 of them were bred individually with normal males (XY) and 17 were identified as XY sex reversed female which produced 67.5% to 85.4% male progeny. Then six identified XY females were mated separately with identified YY males and the produced fry (XY, YY) were feminized with 200mg ẞ-estradiol per kg of feed which produced 92.2% to 98.1% females. These ẞ-estradiol treated females (mixture of XY and YY females) were reared upto maturity and 29 of them were bred individually with normal males (XY) and 8 were identified as YY female. Then three identified putative YY supermales were bred individually with three identified YY females which produced 98.6% to 100.0% male progeny and the resultant progenies were considered as YY supermale broodstock. A total of 21 YY supermales from this broodstock were bred individually with normal females (XX) to produced genetically male tilapia (GMT), 18 of them were identified as putative YY males that produced 81.6% to 100.0% male progeny and the resultant progenies were considered as GMT. The growth, survival and production performance of GMT derived from YY males were compared with sex reversed tilapia (SRT) fry produced through masculinization with 17amethyltestosterone and mixed sex tilapia (MST) in six earthen ponds with a stocking density of 100 fish/decimal. Fry were fed with supplementary feed containing approximately 30% crude protein. The initial weight of GMT, SRT and MST was 5.45±0.35g, 4.35±0.35g and 4.70±0.50g, respectively. Over the culture period of 105 days, the weights of GMT, SRT and MST were found 193.21±3.07g, 176.36±0.94g and 154.82±2.57g respectively and the performances were significantly different (P<0.05) from each other. The survival rate of GMT, SRT and MST was 91.68±1.35%, 87.38±1.38% and 88.16±0.73% respectively and they were not significantly different (P>0.05). The net production of GMT (4252.77±124.06 kg/ha) was significantly (P<0.05) higher than that of SRT (3712.56±71.46 kg/ha) and MST (3269.40±72.01 kg/ha). The result of the economic analysis showed that the net profit of GMT, SRT and MST fish were BDT 0.119±0.007 million, BDT 0.066±0.003 million and BDT 0.029±0.003 million respectively. The net profit of GMT fish was significantly (P<0.05) higher than that of SRT and MST. The development of cryopreservation protocol for YY supermale of Nile tilapia Oreochromis niloticus, entailed a number of experiments. The concentration of sperm ranged from 3.44×109 to 4.40×109 cells/ml. Activation of sperm motility decreased with increasing concentration of the extending media and motility was severely inhibited at 1.0% NaCl. The toxicity of cryoprotectant (DMSO and methanol) to sperm was tested at different concentrations (5%, 10%, 15%) and incubation time (5-20 min). Cryoprotectants with 5% and 10% concentrations produced better motility during 5 and 10 min incubation. Two extenders, Alsever's solution and Modified Fish Ringer, and two cryoprotectants, DMSO and methanol were used for cryopreservation of sperm. Alsever's solution with 10% DMSO showed best performance producing 81.7±1.7% and 53.3±3.3% equilibration and post-thaw motility respectively. Between two diluents, sperm preserved with Alsever's solution plus DMSO produced 36.66±2.32% and 16.20±1.84% fertilization and hatching while fresh sperm yielded 74.34±4.39% and 60.25±5.30% fertilization and hatching rate respectively. The protocol developed through the study can be applied for long-term conservation of genetic materials of sperm of the YY supermales of Nile tilapia Oreochromis niloticus and the cryopreserved sperm can be used in artificial breeding for generating new individuals.