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20213
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Subject: Veterinary Gynaecology and Obstetrics × End date: 2021 × Start date: 2021 × Type: Thesis ×
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    ThesisItemOpen Access
    Study on physico-morphology, biochemical constituents and effect of different levels of egg yolk on cryopreservation of Pantja buck semen
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-09) Rashmi; Gupta, H.P.
    Present study was conducted to assess the cryopreservation of extended Pantja buck semen in Tris-citric acid-fructose-egg yolk-glycerol (TCF-EY) extender containing 4 different levels (5, 10, 15 and 20%) of egg-yolk and to compare the neat, post diluted, post equilibrated and post-thawed quality of semen. Artificial Vagina (AV) method was used to collect semen from four reproductively mature, 2.5 to 4 years old, healthy breeding Pantja bucks. A total of thirty-two ejaculates were collected twice a week. After semen collection from all bucks, neat semen was examined for pH, volume, concentration, mass motility, live spermatozoa count, HOST, acrosome and abnormal spermatozoa percent. Thereafter semen samples were pooled and evaluated for some biochemical parameters viz; Malondialdehyde (MDA), Glutamic Oxaloacetic Transaminase (GOT), Glutamic Pyruvate Transaminase (GPT) and Glutathione Peroxidase (GSH-Px). Pooled semen sample was divided and diluted into four groups as D1[Neat semen + tris-fructosecitric acid-egg yolk (@5%)-glycerol], D2[Neat semen + tris-fructose-citric acid-egg yolk (@10%)-glycerol], D3[Neat semen + tris-fructose-citric acid-egg yolk (@15%)-glycerol] and D4[Neat semen + tris-fructose-citric acid-egg yolk (@20%)-glycerol]. Pooled semen samples were processed and evaluated without washing after dilution, post equilibration and after thawing for progressive motility, live and dead sperm percentage, sperm abnormalities, plasma membrane integrity (HOST) and acrosome integrity. Evaluation of MDA, GOT, GPT and GSH-Px was also done at post thawing stage. Biochemical parameters were evaluated in pooled semen before dilution and at post-thawing stage only. Dilutions were made according to the SOP and equilibrated at 5°C for 3 h and then the French mini (0.25 ml) straws were filled with semen and prefrozen in liquid nitrogen (LN2) vapours followed by stocking in LN2 container for seven days. After 7 days frozen semen straws were thawed in a water bath at 37˚C for 30 seconds and post thaw evaluation was done for above mention parameters. The mean value for neat seminal parameters of Pantja buck semen were recorded as volume 0.51±0.090 ml, pH 6.83±0.004, concentration 3.94±0.228, mass motility 4.49±0.078, live spermatozoa 84.75±1.75%, head abnormality (%) 2.13±0.057 %, mid piece abnormality 0.46±0.021 %, tail abnormality 4.85±0.29%, total abnormality (%) 7.48±0.34 %, plasma membrane integrity (%) 83.53±1.696 %, Intact acrosome (%) 82.63±1.760 %, MDA 2.22±0.16 (nmol/ml), GOT 127.36±0.556 Units/l, GPT 16.99±0.348 Units/l and GSH-Px 7.94±0.56 Units/ml. Significant difference (P< 0.05) was observed for all the parameters in different stages of freezing viz; post dilution, post equilibration and post thawing and in various diluters viz; D1, D2, D3 and D4. All seminal parameters except total abnormality showed higher value in D2 (10% EY) group at post dilution, post equilibration and post thawing stages compared to other groups. Likewise, significantly lower value for MDA, GOT, GPT and GSH-Px was observed in 10% egg yolk at post thawing stage. It was therefore concluded that the Pantja buck semen can be cryopreserved successfully at 10% egg yolk level (V/V) in Tris-citric acid-fructose-egg yolk -glycerol extender.
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    ThesisItemOpen Access
    Insilico characterisation and epitope profiling of differentially expressed plasma membrane proteins of ‘x’ and ‘y’ chromosome bearing bovine sperm
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-09) Sharma, Prachi; Sharma, Mridula
    The specific and differentially expressed plasma membrane proteins of bovine sex sorted semen were selected after reviewing numerous research publications on proteomic profile of sexed semen of bovines. The insilico characters such as physicochemical properties, primary, secondary, tertiary structures, membrane topology, sub-cellular localisation, domain analysis, protein interactions and epitope profiling, both linear and discontinuous peptides was done for all the proteins by using various bioinformatics tools and software. For X specific protein CLRN3, 6 and 4 and for Y chromosome specific protein, SCAMP1, 6 and 8 linear and discontinuous epitopes were predicted, respectively. The proteins were localized on Plasma membrane like SPACA1, Leucine-rich repeat and fibronectin type III domain containing 2, CLRN3, SCAMP1 and Uncharacterised Protein may act as antigenic protein to be used for further production of sex- specific antibodies and separation of X and Y sperm. The results of insilico characterisation and epitope prediction of various selected X and Y bovine sperm plasma membrane specific and differentially expressed proteins would be helpful in selection of some efficient antigenic proteins to be used for further X and Y sperm specific antibody production or synthesis of antibodies artificially.
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    ThesisItemOpen Access
    In silico studies on efficacy of various antibacterial drugs against bovine endometritis
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-09) Arya, Damini; Sharma, Mridula
    The present study was conducted with the objective to find effective drug against bacterial bovine endometritis using Molecular docking. The antibacterial property of selected plants (already studied in vivo and new) was studied its comparison with conventional antibiotics was done. The plants selected for study were Tulsi, Garlic, Aloe vera and Kalmegh, among them Tulsi and Garlic were previously studied in vivo and in vitro for treating bovine endometritis and Aloe vera and Kalmegh were selected on the basis of its potent antibacterial activity. The phytochemicals of Tulsi were thymol, eugenol, linalool, carvacrol, apigenin and caryophyllene. Phytochemicals of Garlic were allicin, ajoene, diallyldisulphide and diallyltrisulphide. Phytochemicals of Aloe vera were aloe-emodin, anthracene, campesterol, lupeol and sitosterol β. Phytochemicals of Kalmegh were andrographolide, 14-deoxyandrographolide, neoandrographolide and 3-O-β-D-Glucopyranosyl-14,19-Dideoxyandrographolide. Selection of bacteria (S. aureus and E. coli) for the present study was done on the basis of most predominantly found bacteria causing bovine endometritis as observed in previous studies. From S. aureus Gyrase B, DNA ligase, dehydrosqualene synthase and FtsA protein and from E. coli DNA gyrase, FtsZ and ZapD protein were selected. The selected bacterial proteins were docked against phytochemicals of plants. To conduct molecular docking the 3-D structure of receptors and ligands is needed. The structures of receptor or bacterial proteins were obtained from Protein Data Bank in PDB format. The 3-D structures of ligand were retrieved from PubChem and ChemSpider databases in SDF and Jsmol format respectively. Based on molecular docking it was concluded that, Aloe vera showed highest binding affinity followed by Kalmegh, Tulsi and Garlic i.e., -8.05, -7.55, -6.40 and -3.89 Kcal/mol respectively. When the antibacterial property of plants and antibiotics were compared it was concluded that the Aloe vera and Kalmegh showed higher binding affinity as compared to gentamicin (-6.95 Kcal/mol) and enrofloxacin (-6.97 Kcal/mol) against pathogenic proteins of Staphylococcus aureus and E. coli. On comparing binding energy obtained by each phytochemical of each plant it was observed that Apigenin of Tulsi, Ajoene of Garlic, Lupeol of Aloe vera and 3-O-β-D-Glucopyranosyl-14,19- Dideoxyandrographolide of Kalmegh showed the better efficacy as they had minimum binding energy against all / maximum bacterial proteins as compared to other phytochemicals of the respective plant. Thus, the combination therapy of Aloe vera, Tulsi and Kalmegh or the specific most efficient phytochemicals (Apigenin, Lupeol, sitosterol β, Anthracene, Neoandrographolide and 3-O-β-DGlucopyranosyl-14, 19-Dideoxyandrographolide) combination may be used for further in vivo or in vitro trials in a direction of effective drug discovery against bacterial bovine endometritis.