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Effect Of EDTA As Chelating Agent In Extender On Post Thaw Quality Of Buffalo Bull Spermatozoa

By: Nadir Hussain (2009-VA-128) | Dr. M. Usman Mehmood.
Contributor(s): Dr. Syed Murtaza Hassan Andrabi | Dr. M. Irfan-ur-Rehman Khan | Prof. Dr. Mansur-ud-Din Ahmad.
Material type: materialTypeLabelBookPublisher: 2016Description: 37p.Subject(s): Department of TheriogenologyDDC classification: 2506-T Dissertation note: Pakistan is an agricultural country where livestock has main contribution in the agriculture. Livestock fulfils the demands of milk, meat and hides. Nili-Ravi is the most important and well developed breed of buffalo and also the highest milk producer among buffalo breeds. Artificial insemination (AI) is the main genetic improvement tool in domestic animals. Spermatozao is cryopreserved for A.I. but is prone to mechanical and biochemical damages during cryopreservation. Numerous strategies have been implemented to overcome this problem which include supplementation of different additives in semen extender to avoid the damages and improve post thaw quality. One of the reason for poor fertility with cryopreserved semen is early capacitation and acrosome reaction signaled by calcium. Early capacitation and acrosome reaction be suppressed by adding calcium chelators. Therefore, present study was aimed to evaluate the effect of EDTA as a calcium chelator in semen extender. The study was conducted at National Agricultural Research Center, Islamabad. Semen was collected twice in a week from 4 mature semen donor Nili-Ravi buffalo bulls maintained at same management conditions. Total of 4 ejaculates were collected and each ejaculate was further divided into 4 aliquots on the basis of EDTA concentration. Group I (0% EDTA), group II (0.1% EDTA), group III (0.2% EDTA) and group IV (0.3% EDTA). Semen was cryopreserved and on post thaw visual motility, acrosome integrity, plasma membrane integrity and chromatin integrity were assessed. Sperm motility and motion characters were assessed through computer assisted semen analyzer (CASA). Through CASA, total motility %, VSL μm/sec (straight line velocity), VCL μm/sec (curve line velocity), VAP μm/sec (average path velocity), BCF (beat cross frequency), LIN % (linearity) and STR % (staginess) were observed. Observations of CASA was were taken at 2 different time intervals i.e. 30 and 60 minutes after post thaw. Data was analyzed by statistical Summary 31 software one way ANOVA (MINITAB v 12.22, State College, Pennsylvania, USA) and was presented as (mean ± SEM). Significant level was set to be (p<0.05). Group III improved visual motility, progressive motility %, plasma membrane integrity significantly (p<0.05). Results of BCF, VCL, VSL and VAP were highest in group III which differed with other three groups significantly (p<0.05). 0.3% EDTA deteriorated most of the parameters of semen observed. Control group and 0.1% behaved almost in similar pattern and no significant different was observed between these two groups but they did not improved all the parameters like group III. hence it can be concluded that 0.2% EDTA improves most of the semen parameters observed so it should be included in semen extender to have better post thaw quality of Nili-Ravi buffalo bull semen.
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Pakistan is an agricultural country where livestock has main contribution in the agriculture. Livestock fulfils the demands of milk, meat and hides. Nili-Ravi is the most important and well developed breed of buffalo and also the highest milk producer among buffalo breeds. Artificial insemination (AI) is the main genetic improvement tool in domestic animals. Spermatozao is cryopreserved for A.I. but is prone to mechanical and biochemical damages during cryopreservation. Numerous strategies have been implemented to overcome this problem which include supplementation of different additives in semen extender to avoid the damages and improve post thaw quality. One of the reason for poor fertility with cryopreserved semen is early capacitation and acrosome reaction signaled by calcium. Early capacitation and acrosome reaction be suppressed by adding calcium chelators. Therefore, present study was aimed to evaluate the effect of EDTA as a calcium chelator in semen extender. The study was conducted at National Agricultural Research Center, Islamabad. Semen was collected twice in a week from 4 mature semen donor Nili-Ravi buffalo bulls maintained at same management conditions. Total of 4 ejaculates were collected and each ejaculate was further divided into 4 aliquots on the basis of EDTA concentration. Group I (0% EDTA), group II (0.1% EDTA), group III (0.2% EDTA) and group IV (0.3% EDTA). Semen was cryopreserved and on post thaw visual motility, acrosome integrity, plasma membrane integrity and chromatin integrity were assessed. Sperm motility and motion characters were assessed through computer assisted semen analyzer (CASA). Through CASA, total motility %, VSL μm/sec (straight line velocity), VCL μm/sec (curve line velocity), VAP μm/sec (average path velocity), BCF (beat cross frequency), LIN % (linearity) and STR % (staginess) were observed. Observations of CASA was were taken at 2 different time intervals i.e. 30 and 60 minutes after post thaw. Data was analyzed by statistical
Summary
31
software one way ANOVA (MINITAB v 12.22, State College, Pennsylvania, USA) and was presented as (mean ± SEM). Significant level was set to be (p<0.05). Group III improved visual motility, progressive motility %, plasma membrane integrity significantly (p<0.05). Results of BCF, VCL, VSL and VAP were highest in group III which differed with other three groups significantly (p<0.05). 0.3% EDTA deteriorated most of the parameters of semen observed. Control group and 0.1% behaved almost in similar pattern and no significant different was observed between these two groups but they did not improved all the parameters like group III. hence it can be concluded that 0.2% EDTA improves most of the semen parameters observed so it should be included in semen extender to have better post thaw quality of Nili-Ravi buffalo bull semen.

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