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20112
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Subject: Animal Reproduction and Gynecology × End date: 2011 × Start date: 2010 ×
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    ThesisItemOpen Access
    FREEZABILITY OF BUCK SPERMATOZOA TREATED WITH CHELATING AGENT FOR FIBRONECTIN TYPE II PROTEINS
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES-MANNUTHY,THRISSUR, 2011) SHINY M.; Hiron M. Harshan
    A study to evaluate the effect of Fn2 type protein chelating agent (choline chloride) or immediate extension and washing on the freezability of buck spermatozoa was carried out at Artificial Insemination Centre, under the department of Animal Reproduction, Gynaecology and Obstetrics, College of Veterinary and Animal Sciences, Mannuthy, Thrissur using 18 normal ejaculates from an adult healthy Malabari crossbred buck. The total seminal plasma protein content was estimated by Lowry’s method. The ejaculates were grouped into three with group I consisting of ejaculates collected directly into Tris extender alone, group II consisting of ejaculates collected into Tris extender containing choline chloride and group III serving as the control. The processed semen was packed in 0.25 ml French straws after extending them in Tris yolk glycerol extender and cryopreservation was carried out. The volume of buck semen samples collected under group III (control) were 0.57 ± 0.09 ml and average sperm concentration was 3971 ± 99.42 million per ml. The gross semen characteristics except volume (0.6 ± 0.09 ml and 0.32 ± 0.03 ml respectively) and sperm concentration (3683 ± 115.49 and 3905 ± 147.50 million per ml) could not be evaluated directly from semen samples of group I and group II as these were collected directly into a collection vial containing Tris extender and choline chloride solution respectively. Mean progressive motile spermatozoa of the three groups were 83.83 ± 1.25, 82.67 ± 1.26 and 84.67 ± 0.84 per cent respectively, while the mean live sperm percentage were observed to be 94.83 ± 0.83, 94.17 ± 1.08, 95.33 ± 0.61 respectively in the three groups. Average per cent of abnormal spermatozoa of the three groups were 3.00 ± 0.63, 3.00 ± 0.68 and 2.50 ± 0.56 respectively. Mean per cent of acrosomal integrity of spermatozoa were 90.50 ± 0.99, 90.17 ± 1.17 and 89.67 ± 0.95 respectively in three groups. After equilibration, the percentage progressive sperm motile spermatozoa in group I, II and III (control) were 76.17 ± 1.08, 77.17 ± 1.08 and 73.67 ± 1.15 respectively. The viable spermatozoa percentage of three groups observed at prefreeze stage were 82.67 ± 1.36, 83.33 ± 0.76 and 79.33 ±0.99 respectively which did not show any significant difference between the groups. The intact acrosome percentage at prefreeze levels were 83.67 ± 1.12 in group I, 82.00 ± 0.97 in group II and 82.67 ± 0.95 in group III. The sperm abnormality of the three groups were 3.33 ± 0.56, 3.33 ± 0.42 and 3.50 ± 0.67 respectively. The mean of membrane integrity by hypo osmotic swelling test of group I, group II and group III were 67.67 ± 2.04, 65.17 ± 2.02, 61.17 ± 1.66 respectively. Spermatozoa of group I had significantly higher HOS response than spermatozoa of group III (control). After freezing and thawing, the mean percentage of progressively motile spermatozoa were found to be 40.83 ± 0.91 in group I, 44.67 ± 1.31 in group II and 33.83 ± 1.30 in group III. The post-thaw live spermatozoa were 49.17 ± 0.91, 52.17 ± 1.30 and 44.17 ± 1.40 respectively. The values obtained in both the treatment groups differed significantly from the values observed in the control group. The values of intact acrosome in percentage were 65.83 ± 1.35 in group I, 71.00 ± 1.00 in group II and 63.33 ± 1.05 in group III. The spermatozoa of choline chloride treated group had significantly higher intact acrosome than spermatozoa of both group I and control group. The percentage of sperm abnormality obtained after thawing were 11.33 ± 0.56 in group I, 10.67 ± 0.49 in group II and 11.17 ± 0.60 group III respectively. The mean of membrane integrity by hypo osmotic swelling test of group I, group II and group III were 43.83 ± 2.50, 48.17 ± 2.30, 37.17 ± 1.89 respectively. Significant difference (p<0.05) was obtained between teh HOS response of spermatozoa of choline chloride treated group and the control group. It was observed that both immediate extension and washing of Malabari buck spermatozoa or treatment with choline chloride at the time of semen collection resulted in improved sperm post thaw characters. Of the two techniques, treating spermatozoa with choline chloride at the time of collection resulted in better post thaw motility, viability acrosome integrity and HOS response of frozen buck semen.
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    ThesisItemOpen Access
    GONADOTROPHINS MEDIATED SUPEROVULATORY RESPONSE AND VIABILITY OF FRESH AND VITRIFIED RABBIT EMBRYOS
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES-MANNUTHY,THRISSUR, 2011) VEENA C. PHILIP; Metilda Joseph
    Superovulation was induced in two groups (A and B) of Newzealand White female rabbits by administration of a single dose of 150 IU of eCG intramuscularly in group A, and two 3 mg and then four 4 mg injections of p-FSH subcutaneously at 12 hour intervals, in group B. On observation of heat signs, mating of females with the same buck was done twice and to induce ovulation 150 IU hCG was given intravenously. Animals belonging to group C remained as control. In vitro collection of embryos were done 72 h post coitum. The onset and intensity of oestrum, number of ovulations, embryo recovery and quality of embryos were studied and compared with those of control animals. Eighteen transferrable embryos from each group were subjected to OPS vitrification. After a minimum storage period of 10 days, the vitrified embryos were thawed and examined for morphological characteristics and membrane integrity was assessed by Trypan blue staining. Further, viability of embryos after in vitro culture was also assessed. The mean interval from gonadotrophin administration to onset of oestrus in groups A and B were 77±5 h and 75±3.30 h, respectively. The intensity of heat was higher in treatment groups than in control. The percentage of animals which showed sexual receptivity in A, B and C groups were 66.67, 100 and 100 respectively. The mean ovulation rate in groups A, B and control were 8.50±5.22, 10.67±3.25 and 7.50±1.38, respectively. The range of corpora lutea was 0 to 33 in group A, 0 to 21 in group B and 2 to 12 in group C. Eventhough total ovulation points were more in group B followed by groups A and C, there was no significant difference in the ovulation rate between the three groups. The total number of haemorrhagic follicles in the groups A, B and control were 1.00±0.63, 10.67±4.64 and 2.00±0.58, respectively. The average number of anovulatory follicles in groups A, B and C were observed as 1.49±0.23, 2.40±0.20 and 1.76±.08, respectively. Significant difference (p<0.05) could be seen in the number of haemorrhagic and anovulatory follicles between group B and other two groups. The mean total ovarian response in the groups A, B and C were 11±6.50, 26.33±7.45 and 11.67±1.9, respectively which did not show any significant difference. 119 The average (percentage) embryo recovery rate in groups A, B and C were 5.50±3.83 (55.56%), 7.83 (66.50%) and 4.83±1.72 (53.41%), respectively. The mean fertilization rate in groups A, B and C were100, 100 and 93.55 per cent, respectively. The corresponding values for mean (percentage) transferrable embryo recovery rate in the three groups were 5.50±3.83 (100), 7.50±2.83 (95.74) and 3.33±1.20 (68.97) respectively. Overall total (percentage) of morulae and blastocysts recovered 72 h post coitum was 49 (44.95) and 60 (55.05) respectively. Percentage embryo recovery rate after OPS vitrification in groups A, B and C were 72, 77.78 and 55.56, respectively. Percentage of morphologically normal embryos recovered from animals belonging to these groups were 92.30, 71.43 and 80, respectively. After Trypan blue staining of morphologically normal embryos, the percentage of viable embryos in the three groups were as 81.8, 83.33 and 71.43, respectively. On further viability assessment of vitrified embryos after TB staining using in vitro culture revealed development of 77.78 per cent of the embryos after 24 h. Present study revealed that administration of gonadotrophins followed by mating twice at the induced heat and hCG supplementation could be effectively used for producing satisfactory superovulatory response in rabbits. The study also indicated that Open pulled straw vitrification technique can be successfully applied for cryopreservation of rabbit embryos. Trypan blue staining technique can be employed as a satisfactory method for assessing the viability of rabbit embryos.