Evaluation of quail and turkey egg yolk for cryopreservation of Nili-Ravi buffalo bull semen.

Egg yolk is used as a cryoprotectant for semen in different mammalian species including buffalo. Egg yolk from different sources may affect freezability of buffalo bull semen. Quail egg yolk (QEY) and turkey egg yolk (TEY) in Tris-citric acid extender was evaluated for post-thaw quality and in vivo fertility rate of cryopreserved buffalo bull semen. Ejaculates were collected on weekly basis from six Nili-Ravi buffalo bulls (12 ejaculates/bull) for a period of 6 weeks and diluted at 37 °C with tris-citric egg yolk extender (50 × 106 motile spermatozoa mL-1) containing different levels of QEY or TEY (5%, 10%, 15%, and 20%) or 20% chicken egg yolk (CEY; controls) and cryopreserved. Percent post-thaw sperm motility (48.3 ± 3.8), plasma membrane integrity (67.9 ± 5.3), live/dead ratio (68.2 ± 5.0), and viability (50.5 ± 3.7) were recorded higher (P < 0.05) in extender containing 5% QEY compared with control. However, TEY at 10% in extender improved (P < 0.05) the post-thaw sperm motility (57.5 ± 5.2), plasma membrane integrity (53.5 ± 4.5), livability (75.3 ± 6.0), and viability (73.5 ± 6.5) compared with higher concentrations of TEY and controls (20% CEY). The chromatin damage (2.0 ± 0.9) and intracellular enzymes, glutamic oxaloacetic transaminase (24.8 ± 3.5) and lactic dehydrogenase (77.7 ± 4.5), release were lower (P < 0.05) in extender containing 10% TEY compared with the controls. Invivo fertility was compared after artificial insemination with semen from two buffalo bulls that was cryopreserved in extenders containing 5% QEY, 10% TEY, or 20% CEY. A total of 600 inseminations (200 inseminations per extender) were recorded; the overall fertility rate was significantly higher (P < 0.05) with semen cryopreserved in extender containing 5% QEY (57.5 vs. 42%) and 10% TEY (57.5 vs. 42%). compared with 20% chiken egg yolk. In conclusion, QEY at 5% and TEY at 10% offers advantages over 20% CEY in terms of in vitro post-thaw semen quality and in vivo fertility of buffalo.

Akhter S ，Rakha BA ，Ansari MS ，Husna AU ，Iqbal S ，Khalid M ... -  《-》

Evaluation of pigeon egg yolk for post thaw quality, enzyme leakage and fertility of buffalo (Bubalus bubalis) bull spermatozoa.

Egg yolk is used as a cryoprotectant in semen preservation. However, its composition varies according to the species which may influence its effectiveness during the freeze-thaw process. Therefore, study was conducted to identify the optimum level of pigeon egg yolk (PEY) in Tris citric acid (TCA) extender for freezability and in vivo fertility of buffalo semen. Semen was collected at weekly intervals for a period of three weeks (replicates) from 6 Nili Ravi buffalo bulls (2 ejaculates/bull/replicate) and diluted with TCA extender (50 × 106 motile spermatozoa ml-1) containing 5%, 10%, 15% and 20% PEY or 20% CEY (control) and cryopreserved. Post-thaw sperm quality and extracellular enzymes leakage was assessed after thawing. Sperm motility, plasma membrane integrity, livability and viability was significantly higher in extenders containing 10% and 15% PEY compared to 5% PEY, 20% PEY or 20% CEY (controls). A dose-dependent decrease was recorded in the chromatin damage for the PEY, being lowest for the 15% and 20% PEY which was significantly less compared to controls (20% CEY). The extracellular GOT and LDH leakage was significantly lower (P < 0.05) in extender containing 10% and 15% PEY compared to the controls. Semen collected from 2 bulls, cryopreserved in extenders containing 15% PEY or 20% chicken egg yolk was assessed for fertility after artificial inseminations. A total of 400 buffaloes were inseminated (100 inseminations/extender/bull). The overall fertility rate was significantly higher (P < 0.05) with semen cryopreserved in extender containing 15% PEY (56%) compared to 20% CEY (42%; controls). In conclusion, pigeon egg yolk at 15% offers advantages over 20% chicken egg yolk in terms of in vitro post-thaw semen quality and in vivo fertility of buffalo.

Akhter S ，Rakha BA ，Ansari MS ，Iqbal S ，Khalid M ... -  《-》

Cryopreservation of Nili-Ravi buffalo (Bubalus bubalis) semen in AndroMed(®) extender; in vitro and in vivo evaluation.

The study was designed to evaluate AndroMed® for the freezability and fertility of Nili-Ravi buffalo semen. Semen was collected from four adult Nili-Ravi buffalo (Bubalus bubalis) bulls for 3 weeks (replicate). Semen ejaculates from each buffalo bull were divided into three aliquots. One aliquot was used for evaluation of motility, plasma membrane integrity, livability, viability, DNA integrity and normal apical ridge. Remaining two aliquots were diluted (37°C; 50 × 106 spermatozoa/ml) in tris-citric egg yolk or AndroMed® extender and cryopreserved in 0.5 ml French straws. After thawing, per cent post-thaw motility (47.9 ± 0.8, 49.2 ± 1.7), plasma membrane integrity (44.4 ± 1.2, 46.8 ± 1.8) and normal apical ridge (81.4 ± 0.3, 83.2 ± 0.3) were recorded similar (p > .05) in tris-citric egg yolk and AndroMed® extender. Higher (p < .05) percentage of sperm livability (70.5 ± 1.4 and 64.4 ± 1.0), viability (67.5 ± 1.5 and 61.5 ± 0.6) and DNA integrity (97.0 ± 0.3 and 93.4 ± 0.21) were recorded in AndroMed® compared to tris-citric egg yolk post-thaw. Values for all the aforementioned spermatozoal quality parameters were observed lower (p < .05) in frozen-thawed compared to fresh semen irrespective of the experimental extenders. Fertility rates of buffalo semen did not differ (p > .05) either cryopreserved in tris-citric egg yolk or AndroMed® extender (45.5% vs. 49%). It is concluded that AndroMed® is capable in protecting the buffalo bull sperm during freeze-thawing process and can be adopted safely for routine use replacing the tris-citric egg yolk extender in artificial insemination programme.

Ansari MS ，Rakha BA ，Akhter S 《-》

Evaluation of duck egg yolk for the cryopreservation of Nili-Ravi buffalo bull semen.

This study was carried out to investigate if the substitution of chicken egg yolk (CEY) with duck egg yolk (DEY) in extenders can improve the quality of frozen-thawed semen of Nili-Ravi buffalo bulls and to study if reducing DEY level in extender affects the freezability results. Thirty semen samples collected from three buffalo bulls were diluted in extenders A, B, C, D and E containing tris, citric acid, fructose, egg yolk, glycerol and antibiotics. Extender A contained 20% CEY (control), while extenders B, C, D and E contained 5, 10, 15 and 20% DEY, respectively. After freezing and storage for 24h in liquid nitrogen, samples were evaluated for post-thaw quality. The post extension sperm motility did not differ between extenders A (control) and E (20% DEY). The same was true for post-thaw percentage of sperm with functional plasma membrane and percentage of sperm with abnormal heads or mid pieces. However, extender E showed higher (P<0.05) values for post-thaw sperm motility, livability and absolute index of livability of spermatozoa at 37 °C compared to extender A. Spermatozoa with abnormal tail were lower (P<0.05) in extender E compared to extender A. Values of these parameters of post-thaw semen quality were highest for extender E containing 20% DEY and decreased significantly with decrease in the concentration of DEY, except sperm abnormalities (head, mid-piece and tail) which increased with decrease in DEY level. These results showed that replacement of 20% CEY with 20% DEY in extenders significantly improved post-thaw sperm motility, livability and absolute index of livability of spermatozoa and reduced tail abnormalities. Reduction in the level of DEY in extenders from 20% adversely affected post-thaw semen quality of Nili-Ravi buffalo bulls.

Waheed S ，Ahmad N ，Najib-ur-Rahman ，Jamil-ur-Rahman H ，Younis M ，Iqbal S ... -  《-》

In vitro evaluation of liquid-stored buffalo semen at 5°C diluted in soya lecithin based extender (Bioxcell®), tris-citric egg yolk, skim milk and egg yolk-citrate extenders.

This study was designed to compare the quality of liquid-stored buffalo bull spermatozoa in soya lecithin based extender Bioxcell(®) (BIOX), milk (MILK), tris-citric egg yolk (TEY) and egg yolk-citrate (EYC) extender at 5°C. Semen was collected from five Nili-Ravi buffalo (Bubalus bubalis) bulls of 6-7 years of age with artificial vagina over a period of 3 weeks (two consecutive ejaculates once in a week). Semen ejaculates having more than 60% motility were pooled, split into four aliquots, diluted (37°C; 10 × 10(6) motile spermatozoa/ml), cooled from 37 to 5°C in 2 h (0.275°C/min) and stored for 5 days. Sperm motility, viability, plasma membrane integrity (PMI) and normal acrosomal ridge were studied at first, third and fifth day of storage. Higher values of progressive sperm motility (%), sperm viability (%), sperm PMI (%) and normal apical ridge (%) were observed in BIOX, MILK and TEY extenders at first, third and fifth day of storage than EYC extender. Progressive sperm motility, sperm viability and sperm PMI in BIOX(®) extender were not different from MILK and TEY extenders at 1st and third day storage period. However, at fifth day of storage, the values for these parameters remained significantly higher (p<0.05) in BIOX(®) compared with MILK, TEY and EYC extenders. At fifth day of storage, the semen quality parameters for Bioxcell(®) were comparable to those with MILK and TEY extenders at third day of storage. In conclusion, motility, viability and PMI of buffalo bull spermatozoa remained similar in Bioxcell(®) , milk and TEY extender at first and third days of storage at 5°C. Yet, the values for the aforementioned parameters in Bioxcell(®) were higher compared with milk, TEY and EYC extender at fifth day of storage at 5°C.

Akhter S ，Ansari MS ，Rakha BA ，Ullah N ，Andrabi SM ，Khalid M ... -  《-》