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Anand Agricultural University, Anand

Anand Agricultural University (AAU) was established in 2004 at Anand with the support of the Government of Gujarat, Act No.(Guj 5 of 2004) dated April 29, 2004. Caved out of the erstwhile Gujarat Agricultural University (GAU), the dream institution of Sardar Vallabhbhai Patel and Dr. K. M. Munshi, the AAU was set up to provide support to the farming community in three facets namely education, research and extension activities in Agriculture, Horticulture Engineering, product Processing and Home Science. At present there seven Colleges, seventeen Research Centers and six Extension Education Institute working in nine districts of Gujarat namely Ahmedabad, Anand, Dahod, Kheda, Panchmahal, Vadodara, Mahisagar, Botad and Chhotaudepur AAU's activities have expanded to span newer commodity sectors such as soil health card, bio-diesel, medicinal plants apart from the mandatory ones like rice, maize, tobacco, vegetable crops, fruit crops, forage crops, animal breeding, nutrition and dairy products etc. the core of AAU's operating philosophy however, continues to create the partnership between the rural people and committed academic as the basic for sustainable rural development. In pursuing its various programmes AAU's overall mission is to promote sustainable growth and economic independence in rural society. AAU aims to do this through education, research and extension education. Thus, AAU works towards the empowerment of the farmers.

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Subject: Veterinary Gynaecology and Obstetrics × End date: 2014 × Start date: 2012 × Type: Thesis × Thesis Type: M.V.Sc. ×
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    ThesisItemOpen Access
    COMPARATIVE EFFICACY OF EGG YOLK AND SOYA BASED EXTENDERS FOR REFRIGERATION (5 °C) AND CRYOPRESERVATION (-196 °C) OF BUFFALO SEMEN
    (AAU, Anand, 2014) Dineshkumar Chaudhary V.; Dr A.J. Dhami
    The present investigation was undertaken during the favourable breeding season (November-February) of the year 2013-2014 on six mature Surti buffalo bulls at Central Sperm Station of Department of Gynaecology and Obstetrics of the Veterinary College, AAU, Anand. The study covered evaluation of seminal characteristics in neat semen and then comparative efficacy of egg yolk based standard TFYG (Tris-citric acid-fructoseegg yolk-glycerol) extender and soybean based commercially available extenders (Bioxcell® and Optixcell®, IMV, France) using split-ejaculate technique through various morphological and functional attributes of spermatozoa extended/preserved/ processed in these extenders for refrigeration preservation (at 5°C up to 72 hrs) and cryopreservation (-196°C), including interrelationships of quality sperm parameters of fresh, refrigerated and cryopreserved semen. Immediately after collection, the ejaculates (8 per bull) were evaluated for routine physico-morphological attributes, including motility, viability, morphology (eosinnigrosin), acrosomal integrity (Giemsa stain) and plasma membrane integrity (HOST 150 mOsm/L; an in vitro fertility test) through standard procedures and using phase contrast microscope. The ejaculates were divided in to three equal aliquots, and extended at the concentration of 100 ×106 spermatozoa ml-1 at 34°C with 3 different extenders. Small portions of the extended semen samples (2 ml from each aliquot) were transferred to a refrigerator for 5°C preservation and evaluated at 24 hrs interval up to 72 hrs for above quality parameters. The remaining portions of extended semen samples were used for filling the French mini straws on IS4 system (IMV, France). After gradual cooling over 60-90 minutes and equilibration for 4 hrs in cold handling cabinet, the straws were frozen in liquid nitrogen vapour using a programmable bio-freezer (Digitcool 5300 CE ZH 350, IMV, France). The straws of all three extenders were evaluated at pre-freezing (after equilibration) and after 24 hrs of freezing (post-freeze stage) for the above quality parameters. Post-thaw incubation test (37 ºC) was also performed to evaluate sperm survival at 30 and 60 min of incubation. The mean values of ejaculate volume, density (1-4 score), sperm concentration, mass activity (0-5 score), individual sperm motility, live sperm, intact acrosome and HOS reactive sperms observed in fresh semen were 3.43±0.10 ml, 2.14±0.11, 744.90± 36.38 million/ml, 3.45±0.07, 78.54±0.51 per cent, 90.48±0.33 per cent, 94.40±0.20 per cent and 79.35±0.42 per cent, respectively. The variation between bulls was significant (P<0.05) for most of these traits. The overall segment-wise per cent abnormalities of sperm head, midpiece and tail region recorded in fresh semen were 1.83±0.09 0.88±0.06 and 3.44±0.08, respectively, with the total of 6.15±0.15 per cent. The mean percentages spermatozoa having swollen, ruffled, detached and denuded acrosome in fresh semen were found to be 2.29±0.08, 1.60±0.10, 1.10±0.09 and 0.60±0.08, respectively. The overall pooled mean percentages of progressively motile spermatozoa (irrespective of extenders) observed on dilution (0-hr), and after 24, 48, 72 hrs of refrigeration preservation of semen at 5°C were 78.54±0.30, 68.30±0.30, 59.38±0.35 and 51.63±0.47. The corresponding values for live sperm per cent were 90.48±0.19, 76.89± 0.51, 69.51±0.35 and 61.86±0.45; those of morphologically normal sperm per cent 93.85±0.08, 90.70±0.11, 89.08±0.10 and 87.66±0.10; intact acrosome per cent 94.40± 0.12, 88.22±0.13, 84.60±0.14 and 81.70±0.17, and HOS reactive sperm (intact plasma membrane) per cent 79.35±0.24, 65.99±0.31, 57.69±0.36 and 50.70±0.43, respectively, all of which differed highly significantly (P<0.01) between storage intervals. The mean percentages of progressively motile spermatozoa observed in Optixcell, TFYG and Bioxcell extenders at 24, 48, 72 hrs of refrigeration storage were 70.31±0.46, 68.33±0.45, 66.25±0.46; 61.46±0.51, 59.79±0.58, 56.88±0.55, and 53.85± 0.75, 52.50±0.68, 48.54±0.84, respectively. The corresponding values for live sperm per cent in Optixcell, TFYG and Bioxcell extenders at three intervals were 78.35±1.36, 77.35±0.46, 74.96±0.47; 71.96±0.48, 69.90±0.54, 66.67±0.54, and 64.38±0.70, 62.56± 0.65, 58.65±0.78, and those of morphologically normal sperms were 91.37±0.16, 90.65±0.15, 89.79±0.14; 90.08±0.15, 88.20±0.11, 88.25±0.12, and 88.54±0.12, 87.35± 0.14, 87.08± 0.15, respectively. The mean percentages of spermatozoa with intact acrosomes observed in Optixcell, TFYG and Bioxcell extenders at 24, 48, 72 hrs of refrigeration storage were 89.27±0.20, 88.15±0.18, 87.25±0.21; 85.58±0.24, 84.63± 0.18, 83.60±0.24, and 82.83± 0.27, 81.67±0.23, 80.60±0.30, respectively. The corresponding values for sperms with intact plasma membrane (HOS reactive sperms) in Optixcell, TFYG and Bioxcell extenders at three intervals were 68.10±0.46, 66.04±0.50, 63.81±0.45; 60.00±0.55, 57.92±0.55, 55.15±0.56, and 53.40±0.68, 51.35±0.60, 47.35±0.68, respectively. The results were significantly (P<0.05) superior with Optixcell followed by TFYG extender than with the Bioxcell. There were gradual and significant (P<0.05) decline in progressively motile, viable, morphologically normal, acrosomal intact and plasma membrane intact sperm per cent in all the three extenders with each increase in 24 hourly storage interval at 5°C. The overall pooled mean percentages of progressively motile spermatozoa observed (irrespective of extenders) at initial, pre-freeze and post-thaw stage of buffalo semen were 78.54±0.30, 69.48±0.26, and 47.33±0.52. The corresponding values for live sperm per cent were 90.48±0.19, 79.39±0.28 and 56.90±0.52; those of morphologically normal sperm per cent 93.85±0.08, 92.14±0.10 and 87.88±0.12; intact acrosome per cent 94.40±0.12, 89.55±0.14 and 77.08±0.17, and HOS reactive sperm (intact plasma membrane) per cent 79.35±0.24, 67.94±0.26 and 45.05±0.52, respectively, all of which differed significantly (P<0.01) between freezing stages. The overall mean post-thaw incubation (37°C) survival of spermatozoa immediately after thawing (0-min), and after 30- and 60-min of post-thaw incubation was 47.33±0.85, 41.67±0.85 and 34.62±0.82 per cent (P<0.01), respectively. The mean percentages of progressively motile spermatozoa observed in Optixcell, TFYG and Bioxcell extenders at pre-freeze and post-thaw stages were 70.94± 0.38, 69.48±0.37, 68.02±0.49, and 49.90±0.90, 47.71±0.79, 44.38±0.85, respectively. The corresponding values for live sperm per cent in Optixcell, TFYG and Bioxcell extenders at pre- and post-freeze stage were 81.58±0.38, 79.21±0.39, 77.38±0.48, and 59.67±0.91, 57.19±0.79, 53.85±0.84; and those of morphologically normal sperms 92.92±0.15, 92.10±0.14, 91.40±0.16, and 88.73±0.18, 87.67±0.17, 87.25±0.21, respectively. The mean percentages of spermatozoa with intact acrosomes observed in Optixcell, TFYG and Bioxcell extenders at pre-freeze and post-thaw stage were 90.52± 0.21, 89.54±0.18, 88.58±0.22, and 78.50±0.25, 76.83±0.23, 75.90±0.27, respectively. The corresponding values for sperms with intact plasma membrane (HOS reactive sperms) in Optixcell, TFYG and Bioxcell extenders at pre-freeze and post-thaw stage were 70.23±0.37, 67.96±0.32, 65.65±0.42, and 47.81±0.90, 45.02±0.84 and 42.31±0.82, respectively. The results were significantly (P<0.05) superior in Optixcell followed by TFYG extender as compared to the Bioxcell extender. The mean percentages of progressively motile spermatozoa observed immediate after thawing in TFYG, Bioxcell and Optixcell extenders were 47.71±0.79, 44.38±0.85 and 49.90±0.90, respectively. The corresponding motility values after 30-min of postthaw incubation were 41.98±0.80, 38.44±0.82 and 44.58±0.93 per cent, and those of 60- min of incubation were 35.52±0.79, 31.15±0.85 and 37.19±0.81 per cent, respectively. The results showed that the commercial soya based Optixcell as well as standard egg yolk based TFYG extenders could sustain better and acceptable level of sperm survival at least for 1-hr after thawing. The overall mean percentages of sperms with abnormal head, mid-piece and tail and those with different forms of acrosomal defects like swollen, ruffled, detached and denuded acrosome also followed the pattern of morphologically normal sperm and intact acrosomes during different intervals of refrigeration as well as cryopreservation of extended buffalo semen in different extenders. The values of all these traits differed significantly (P < 0.05) between storage intervals, and increased with increase in storage duration at 5°C or in post-thawed over pre-freeze stage. The sperm protective ability of Optixcell extender was superior followed by TFYG and the least of Bioxcell. The pre-freeze and post-thaw sperm motility, viability, morphology, acrosomal integrity and plasma membrane integrity recorded in all three extenders followed the same trend of declining in each extender at post-thaw over pre-freeze stage. Optixcell was significantly superior, and at par with TFYG, than the Bioxcell in maintaining greater motility, viability, morphology, acrosomal / plasma membrane integrity of buffalo spermatozoa during cryopreservation process as well as in terms of keeping quality at refrigeration (5°C) preservation also. There were significant (P<0.01) interrelationships between sperm motility, viability, normal morphology, intact acrosome and plasma membrane integrity in fresh, refrigerated and cryopreserved buffalo semen, which proved that initial motility and membrane integrity can be used as predicative measures in routine semen evaluation.