Aquaculture Canada and WAS North America 2022

August 15 - 18, 2022

St Johns, Newfoundland, Canada

SEMEN CHARACTERISTICS AND OPTIMIZATION OF SPERM CRYOPRESERVATION PROTOCOL OF Labeo gonius (HAMILTON, 1822)

Hayin Tamut, Raghvendra Singh*, Santosh Kumar, Aditya Kumar, Ajay Kumar Singh, Achal Singh, Ramasankar Sah, Chandrabhushan Nishad, Arvind Verma and Kuldeep K Lal

ICAR-National Bureau of Fish Genetic Resources, canal ring road, PO Dilkusha, Lucknow, 226 002

*corresponding author- email: shakya.raghav@gmail.com

 



Abstract

 The study describe the semen characteristics and experimental results to determine suitable protocol for sperm cryopreservation of minor carp, Labeo

gonius (Kuria

Labeo ), which can be used in seed production. The mean pH of seminal plasma was 7.6 ±0.19, mean osmolality of milt was 276±15 mOsmol kg- 1 and sperm count in milt samples from different males (n=9) were found in the range of 25.4×109 to 34.2×109 cells ml-1. To determine the suitable extenders for cryopreservation a total of 9 different extenders were screened. Finally, three extenders (8, 9B and 9C) having mean sperm activation with water above 70% and mean spermatozoa survival above time ( 40 seconds) were selected for cryopreservation work. Milt to diluent ratio of 1:6 was kept for cryopreservation work.  DMSO (10% v/v) was added in extenders as cryoprotectant to defend from stress on cells during cooling and freezing and diluted milt was then filled in 0.25 ml capacity straw and stored in liquid nitrogen after 20 minutes equilibration time. The feasibility of selected extender and cryoprotectant combinations was assessed by the ability of cryopreserved sperm to fertilize the Labeo

gonius

 eggs and hatching percentage. Fertilized eggs were incubated in flow-through incubation system with a water flow rate of 2-3 litre per minute. Recorded hatching rates and survival rates in extender 8, 9B and 9C were 61.1± 0.23%, 81.3 ± 0.19%, 83.7 ±2.7% and 44.6 ±1.2 %, 45.0 ±2.64% and 46.7± 1.52%, respectively. Control group (having fertilized from fresh sperms) yielded 84.99±0.88% hatching rate and 48.6± 1.52% survival rate. Statistical analysis indicated that the extender 9C and 9B with 10% DMSO gave higher hatching and survival rate in comparison to extender 8 which might be due to the presence of sugar in 9C and 9B. The fertilization capacity of cryopreserved sperms and control group (having fresh milt) were compared and significant difference was not found in terms of hatching rate (p

0.05) as well as for survival rate (p≥ 0.05) with control except for extender 8 which was even higher than 50 percent.  Therefore, extender 8, 9B and 9C were recommended for cryopreservation of milt of  L. gonius which would be helpful in ex-situ conservation of the species.

Keyword s: Labeo

gonius ,  sperm cryopreservation, DMSO, ex-situ conservation.