Optimization of Nile tilapia artificial breeding using Human Chorionic Gonadotropin (hCG) hormone

Global production of Nile tilapia (Oreochromis niloticus) could reach 7.3 million tons by 2030, increasing the pressure to improve efficiency in fry production. To meet this growing need, fish farmers turn to artificial breeding techniques that allow for synchronized spawning and maximized fry production.

One of the most common tools is the use of the human chorionic gonadotropin (hCG) hormone. However, despite its popularity, a detailed and optimized protocol specifically for Nile tilapia was lacking. A study published in the journal Methods and Protocols by Golam Rbbani and a team of researchers from Nord University, GenoMar Genetics, the European University at Saint Petersburg, and the Institute of Marine Sciences (ICM-CSIC) fills this gap, presenting an artificial breeding protocol that is not only effective but also achieves outstanding results with a significantly low hormonal dose.

The challenge: Synchronizing nature for efficient production

Although tilapia is prolific, its reproduction on farms can be asynchronous, influenced by factors such as temperature, photoperiod, or oxygen availability. This variability complicates hatchery management and production planning.

Hormonal induction emerges as a solution to control the process, allowing producers to obtain high-quality gametes predictably. This is not only vital for commercial production but also for genetic improvement programs and advanced scientific studies.

The optimized step-by-step protocol

The study’s objective was clear: to establish an hCG induction method that would optimize fertilization, hatching, and survival rates for aquaculture production and research. The methodology, though rigorous, is based on clear and replicable steps.

Broodstock selection and hormonal dosing

The first step is the correct selection of broodstock. Mature females are identified by characteristics such as a reddish genital papilla and a swollen abdomen. Once selected, the core of the protocol is the istration of a single intramuscular injection of hCG at a dosage of 2 IU/g of body weight. This detail is crucial, as other studies had reported using doses up to 25 times higher to achieve comparable results.

Management and gamete collection

After the injection, the female is kept for 24 hours in a recovery tank with constant aeration and a stable temperature of 28°C. After this period, the eggs are collected by applying a gentle massage to the female’s abdomen. Immediately, sperm is stripped from a mature male and mixed directly with the eggs, adding a small amount of water to activate the sperm and ensure fertilization.

Incubation that mimics nature

The fertilized eggs are washed to remove debris and transferred to ‘egg rocker’ type incubators. This system keeps the eggs in constant motion, simulating the natural mouthbrooding process of female tilapia and preventing them from settling at the bottom. Throughout the process, it is vital to remove dead eggs to avoid compromising water quality.

Key results: High efficacy and survival

The protocol proved to be a resounding success, achieving an average fertilization rate of 88.3% and an impressive larval survival rate of 90.5%. To assess the method’s robustness, the researchers tested incubation at three different temperatures (24°C, 28°C, and 32°C), reflecting the species’ natural thermal range.

• Fertilization: Fertilization rates remained consistently high across all temperatures, ranging from 82.6% to 91.3%.
• Hatching: The hatching rate showed greater variation with temperature, with the highest peak (80.5%) observed in one of the replicates at 24°C.
• Survival: Larval survival was remarkably high and stable across all conditions, with values reaching up to 94.6%.

The study concluded that a temperature of 28°C offered the most balanced results overall, underscoring the importance of maintaining optimal thermal conditions to maximize reproductive efficiency.

Implications for the fish farmer and researcher

Beyond the excellent results, this protocol has two practical implications of great value for the aquaculture sector.

A boost for commercial production

For a tilapia farmer, this method offers a reliable tool to synchronize spawning, ensuring a constant and predictable supply of high-quality larvae. The low hormonal dose not only reduces costs but also minimizes potential residual effects, aligning with more sustainable and efficient production.

An open door to advanced genetic improvement

The protocol is especially relevant for the field of research and genetic improvement. Techniques like gene editing with CRISPR/Cas9 require embryos of very uniform quality and developmental stage to be successful. Traditional egg collection methods often result in uneven batches, which reduces the effectiveness of these technologies. By ensuring the collection of fully mature and synchronized oocytes, this protocol facilitates the microinjection of genetic material and dramatically increases the precision and reproducibility of gene-editing experiments in tilapia.

Conclusion

The study by Rbbani and his team does not simply present another method, but an optimized, validated, and detailed protocol that addresses a specific need in Nile tilapia aquaculture. By achieving high fertilization and survival rates with a reduced hormonal dose, this research offers producers a more efficient and economical strategy for artificial breeding. At the same time, it provides the scientific community with a robust tool to advance reproductive biology and genetic improvement, consolidating tilapia’s role not only as a pillar of aquaculture production but also as a valuable study model.

Reference (open access)
Rbbani, G.; Siriyappagouder, P.; Murshed, R.; Joshi, R.; Nedoluzhko, A.; Galindo-Villegas, J.; Fernandes, J.M.O. Optimization of Nile Tilapia Artificial Breeding Using Human Chorionic Gonadotropin (hCG) Hormone. Methods Protoc. 2025, 8, 57. https://doi.org/10.3390/mps8030057