**2.2 Selection of broodstock**

308 Current Frontiers in Cryopreservation

ringer and freezing in 1-milliliter cryovials in a programmable freezer. To avoid an excess of spermatozoa per egg, post-thaw semen was diluted 1:20, 1:200 or 1:2000 before fertilization. Even frozen- thawed spermatozoa with low numbers of live cells yielded adequate hatching rates. They found out that the maximum sperm dilution ratio to achieve hatching rates similar to control was 1:200 without losing fertilization ability. However, at 1:2000 the hatching rates produced with frozen spermatozoa were lower than the control African catfish. Similarly *Heterobranchus longifilis* spermatozoa were diluted 1:3 before freezing and 1:10 after thawing and had the same fertilization ability (78.9%) as the control (81.1%). On the contrary for *Cyprinus carpio*, no spermatozoa survived when diluted higher than 1: 5

Cryopreservation of catfish spermatozoa is useful as a routine method of gamete storage and management. However, the economic factor should also be considered. The technology of cryopreservation with the use of liquid nitrogen though desirable but is cost intensive. Therefore there is a need to study how the cryopreserved semen will be maximally utilized with good fertilizing results at the same time cost-effective and affordable for the farmers

To avoid wastage of cryopreserved spermatozoa per egg clutch after dilution with physiological salt solution, fertilization of various measures of egg clutches were tested in the present research with differently cryopreserved spermatozoa for optimization and for cost evaluation. In a second study the concentration of the semen was reduced, *i.e*., diluted at a dilution ratio of 1:20 and 1:200 to verify the spermatozoa are not in excess and

Another study was carried out in order to assist the farmers to determine the approximate amount of egg clutch that will be adequate for a milliliter (ml) of cryopreserved semen without wasting spermatozoa in order to evaluate economic cost and profitability. The aim of this study was to verify the possibility of cryopreserving African catfish under long-term condition in liquid Nitrogen (LN2) and evaluate the viability and fertility optimization of a specific amount (*i.e.,* 1 ml ) of cryopreserved semen of African catfish cryopreserved in LN2 using various cryoprotective agents with different measures of egg clutches. This paper aimed to establish a standard fertility ratio between a ml of semen and clutch of eggs in order to prevent wastage of semen; be able to maximize the resources and evaluate profitability of cryopreservation in liquid nitrogen by evaluating the effects on the cryopreserved semen as to motility and hatchability, the ability to hatch the eggs from a

For the benefit of prospective users of cryopreservation of African catfish semen, the whole

The broodstock of the African catfish, *C. gariepinus* were obtained from reliable farms in Ile-Ife and Ibadan, Nigeria and were transported in 25 litre tank opened at the top to the Wet Laboratory of the Department of Animal Sciences, Obafemi Awolowo University, Ile Ife.

before and after freezing (Lubzens *et al.*, 1997).

gravid female and survival of ensuing larvae.

process is pictorially presented in Figures 1-12.

**2. Materials and methods** 

**2.1 Husbandry of the broodstock** 

consequently be wasted.

The sexually matured female was selected according to their swollen, reddish genital papilla and a well distended, swollen soft abdomen. A slight pressure was applied on the abdomen towards the genital papilla after which ripe eggs oozed out which were green-brownish in color and ripe eggs are generally uniform in size. The female broodstock was stocked in the hatchery for about 2 days without feeding so that the alimentary tract was empty at the time of stripping. It is very important that the collected eggs did not get contaminated. Sexually matured male broodstock was selected based on a reddish or pinkish pointed and vascularized genital papilla. The temperature of the broodstock kept in the tank was maintained at 25–270C.
