Aquaculture International 12: 47–56, 2004. © 2004 Kluwer Academic Publishers. Printed in the Netherlands. Successful hybridization of Acipenser species using cryopreserved sperm BÉLA URBÁNYI 1,* , ÁKOS HORVÁTH 1 and BALÁZS KOVÁCS 2 1 Department of Fish Culture, Faculty of Agriculture and Environmental Sciences, Szent Istvan University, Páter K. u. 1., Gödöll˝ o, H-2103, Hungary; 2 Agricultural Biotechnology Center, Szent-Györgyi Albert ut 4, Gödöll˝ o, H-2100, Hungary; * Author for correspondence (e-mail: urbanyib@nt.ktg.gau.hu; phone: +36-28-522-000/1657; fax: +36-28-410-804) Received 10 December 2002; accepted 20 June 2003 Abstract. Successful hybridization of sterlet (Acipenser ruthenus) × Siberian sturgeon (Acipenser baeri), sterlet × Russian sturgeon (Acipenser gueldenstaedti) and sterlet × European sturgeon (Acipenser sturio) was carried out for the first time using cryopreserved semen of sturgeon males and sterlet × sterlet crosses as controls. Sperm of all three species was diluted with a cryodiluent composed of 23.4 mM sucrose, 0.25 mM KCl, 30 mM Tris (pH 8.0) and 10% methanol. The samples were frozen in plastic straws in the vapor of li- quid nitrogen at the height of 3 cm above the level of nitrogen for 3 min. Following thawing approximately 3000 sterlet eggs were fertilized with six straws of frozen-thawed sperm. The hatching rate with sterlet sperm was 30.6% while the hatching rate of A. ruthenus × A. baeri, A. ruthenus × A. gueldenstaedti and A. ruthenus × A. sturio hybrids was 50, 17.4 and 34%, respectively. Morphometric markers as well as random amplified polymorphic DNA (RAPD) assay was used to verify interspecific hybridization. Key words: Cryopreservation, Hybrids, RAPD, Sperm, Sturgeon Introduction Sturgeons (species of the order Acipenseriformes) are chondrostean fishes of ancient origin (Birstein and DeSalle, 1998). Several species are restricted to very small populations and in some cases are close to extinction. Some attempts have been made to understand the problems of sturgeon conservation in order to increase genetic diversity and the chance for their survival (Billard and Lecointre, 2001). The first attempts to cryopreserve sturgeon sperm can be dated back as far as the 1960s, when researchers in the former Soviet Union carried out experiments on this subject (for review see Dettlaff et al., 1993). Since then several protocols have been developed (Drokin et al., 1991; Cherepanov et al., 1993; Drokin et al., 1993; Ciereszko et al., 1996; Tsvetkova et al., 1996; Jähnichen et al., 1999a), but most reported experiments shared one common