Fertilization by intracytoplasmic sperm injection in Nile tilapia (Oreochromis niloticus) eggs

In this report we describe the steps used for the production of Nile tilapia by intracytoplasmic sperm injection (ICSI). To assure a constant and reliable source of eggs for ICSI we used a method where female Nile tilapia, Oreochromis niloticus, spawned when held individually in 80-l tanks (a single-breeding system). This allowed collection of eggs every 28 ± 9 days (mean ± S.D.) from 26 spawning events by 7 females that spawned two or more times. It was common to obtain more than 1000 eggs from a single stripping. The second step in this project involved the extension of egg viability to assure sufficient time for the ICSI procedure. This was accomplished by placing the eggs in Hanks' balanced salt solution after stripping, which extended the period of fertility for at least 3 h after collection. The third step was to minimize chromosomal damage after ICSI by localization of the metaphase plate within eggs. The maternal chromosomes were found to be ∼43 μm from the micropyle (injection site) at the moment of fertilization. The final step was to apply these methods to evaluate the ICSI procedure for tilapia. From a total of 113 Nile tilapia eggs injected with fresh sperm, 7 (5%) were fertilized, 5 (4%) developed abnormally to neurula and 1 (1%) developed normally and reached adulthood. These results demonstrated for the first time that the ICSI procedures here described allow fertilization and subsequent development of Nile tilapia eggs into normal larvae, hatching and beyond. This provides opportunities for the study of basic processes involved in fertilization and zygotic development and expands the utility and range of sperm storage methods such as cryopreservation or freeze-drying.