A sperm cryopreservation protocol for the loach Misgurnus anguillicaudatus and its applicability for other related species

• Published in: Animal reproduction science 2009-12, Vol.116 (3), p.335-345
• Main Authors: Yasui, George Shigueki, Arias-Rodriguez, Lenin, Fujimoto, Takafumi, Arai, Katsutoshi
• Format: Article
• Language: English
• Subjects: Animals; Barbatula toni; Cryopreservation; Cryopreservation - methods; Cryoprotective Agents - chemistry; Cryoprotective Agents - pharmacology; Cypriniformes - physiology; Female; Fertilization - drug effects; Fertilization - physiology; Fish; Fishes - physiology; Freshwater; Inseminating dose; Insemination, Artificial - methods; Lefua; Loach; Male; Misgurnus; Misgurnus anguillicaudatus; Misgurnus mizolepis; Semen Preservation - methods; Sperm; Temperature
• ISSN: 0378-4320; 1873-2232
• DOI: 10.1016/j.anireprosci.2009.02.021

The aim of the present study was to establish a protocol of sperm cryopreservation in Misgurnus anguillicaudatus and verify the applicability of the obtained protocol in other loach species. We evaluated the following parameters: inseminating dose, thawing temperatures (20, 25 and 30 °C for 10 s), extenders (loach or cyprinid extenders), internal cryoprotectants (dimethyl sulfoxide (DMSO), dimethylacetamide (DMA), glycerol (Gly), ethylene glycol (EG), and methanol (MeOH) at 0, 5, 10 and 15%), external cryoprotectants (bovine serum albumin 1 and 2%; sucrose 0.5 and 1%; glucose 0.5 and 1%; glycine 0.5 and 1%), activating solutions (distilled water, dechlorinated tap water, 25 mM NaCl and 50 mM NaCl), and hatchability of the eggs when fertilized with fresh or cryopreserved sperm. After the evaluation of these parameters, we optimized the cryopreservation using the following procedure: thawing temperature at 25 °C for 10 s; loach or cyprinid extenders; methanol at 10 or 15% as internal cryoprotectants; glycine 0.5% or bovine serum albumin 1% as external cryoprotectants and 50 mM NaCl for sperm activation. Using this procedure, the fertilizability of the post-thawed sperm was 47% in comparison to the fresh sperm, at the minimum inseminating dose (687.65 spermatozoa egg −1 mL −1). Based on this protocol, sperm from other loach species Lefua nikkonis, Misgurnus mizolepis and Barbatula toni were cryopreserved successfully.