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ThesisItem Open Access Isolation, characterization and morpho-biometric evaluation of pre-pubertal porcine spermatogonial stem cells in different culture media(College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022-03) Lalmasawma, Timothy; Das, ArpanaTestes samples were collected from 7-15 days old pre-pubertal male crossbred piglets (Local × Hampshire) for isolation, enrichment and in vitro culture of porcine spermatogonial stem cells (SSCs). Isolation of spermatogonial stem cells like cells was performed by double enzymatic digestion using four enzymes viz., collagenase, DNase I, hyaluronidase type II and trypsin-EDTA. Isolated cells were further enriched by differential plating and percoll density gradient centrifugation method. Enriched cells were cultured on Sertoli cell feeder layer in three different culture media. All the three media consisted of same concentration of DMEM, NEEA, L-glutamine, FBS, EGF and FGF, however in addition to these, LIF was added to media I, GDNF was added to media II and both LIF and GDNF were added to media III. Characterization of SSCs was done by alkaline phosphatase and immunoflourescence staining. Expression of SSC specific pluripotent marker genes by putative SSCs was also studied by RT-PCR study. Porcine SSCs were observed as dome shaped round or oval bodies on 5th -6th day of culture in all the three media. Clustering of cell was observed from 4th -5th day of culture and single, paired or multiple colonies were observed from 8-10th day of culture. The SSCs colonies appeared as mulberry, grape or rosette shaped with irregular distinct boundary from feeder layer on 15th - 19th day of culture in all the three media. However, the shape of the SSCs was found to be distorted with increase in the days of culture. The morphology of the SSC colonies was maintained best up to 30th day of culture in media III. The SSC colony number was recorded as 82.14 ± 2.91, 60.07 ± 2. 78 and 48.43 ± 1.96 on 5th, 15th and 30th day of culture respectively in media I. The corresponding numbers were 91.71 ± 2.62, 67.00 ± 2.05 and 57.29 ± 2.17 in media II and 105.93 ± 2.82, 80.21 ± 2.45 and 62.50 ± 2.09 in media III respectively. The SSC colony diameter was found to be 64.26 ± 0.85, 125.30 ± 1.88 and 123.01 ± 5.49μm on 5th, 15th and 30th day of culture respectively in media I. The corresponding values were 69.67 ± 1.12, 139.58 ± 3.93 and 142.08 ± 5.72μm in media II and 76.49 ± 1.61, 152.55 ± 4.07 and 172. 08 ± 4.96μm in media III respectively. The day of culture and culture media had significant effect (P≥0.01) on SSC colony number and significantly higher number of SSC colony was observed on day 5 and lower was on day 30 of culture in all the three media. The SSC colony number was significantly higher in media III containing both GDNF and LIF. The diameter of SSC colony differed significantly (P≥0.05) due to day of culture and culture media. The interaction between day of culture and culture media was also significant (P≥0.01). The colony diameter recorded on day 30 of culture was significantly higher, whereas lower number was recorded on day 5 of culture in all the culture media. Diameter of SSC colony obtained in media III was found to significantly higher and the lower diameter was obtained in media I on all the day of culture. It was observed that the SSC colony number decreased and colony diameter increased with the day of culture from day 5th to 30th day of culture in all the media.The putative SSCs in all the three media showed positive result for alkaline phosphatase and immunofluorescence staining. The putative SSCs in all the three media were also found to express SSC specific pluripotent marker genes viz., OCT4, SOX2, NANOG and maximum expression was observed in media III, however, no expression was recorded for c-KIT and PPARγ which were known to be the markers for differentiated SSCs. BAX4, an apoptopic marker gene was also expressed by putative SSCs in all the three media. Based on the findings of the present study, it may be concluded that a pure population of porcine spermatogonial stem cells (SSCs) could be obtained and successfully maintained in vitro up to 30th day of culture. Media III containing DMEM, FBS, NEAA, L-glutamin, FGF, EGF, LIF and GDNF was found to be the best for in vitro culture of porcine SSCs.ThesisItem Open Access Phenotypic characterization and polymorphism study of prolactin gene in native geese of Assam(2022) Gogoi, Ankita; Das, BulaData pertaining to 1015 native geese of Assam were considered for morphometric characteristics, productive and reproductive performances, egg quality and carcass traits, along with screening of polymorphisms in 5'flanking region of Prolactin gene. Two plumage colours, cinnamon (62.38%) and white (37.62%) were seen. Bill colour was observed to be black (49.08%), orange (35.33%,), yellow (13.24%) and mixture of black and orange (2.35%). The skin was 100% white. Shank and feet colour were orange (75.32%), yellow (21.67%) and mixture of black and orange colour (3.01%). Black (60.14%), brown (28.49%) and blue (11.37%) eye colour was seen. The overall LSM ± SE for body weights at hatching, 4 week, 8 week, 6 month, 8 month,12 month of age were 89.85±0.11, 66.89±1.21, 1761.90±2.08, 3305.42±4.63, 3575.80±10.61 and 3804.84±3.91 g, respectively. The overall mean (cm) for bill length, bill width, knob diameter (at 12 months of age), head length, head width, neck length, neck girth, breast length, keel length, body length, body circumference, shank length and wingspan (3 to 6, 8 to 10 and 12 months of age) at the respective ages were 5.31 ± 0.018, 6.24 ± 0.01, 7.24 ± 0.012, 8.42 ± 0.014; 1.46 ± 0.010, 1.95 ± 0.008, 2.19 ± 0.009, 2.45 ± 0.009; 2.34 ± 0.006; 5.42± 0.019, 6.70 ± 0.018, 8.31 ± 0.019, 10.26 ± 0.018; 2.44 ± 0.009, 2.80 ± 0.011, 3.03 ± 0.010, 3.26 ± 0.012; 14.13 ± 0.031, 16.00 ± 0.032, 17.80 ± 0.026, 21.74 ± 0.027; 2.17 ± 0.012, 4.12 ± 0.013, 6.28 ± 0.011, 10.19 ± 0.019; 7.95 ± 0.022, 15.73 ± 0.044, 21.42 ± 0.023, 29.93 ± 0.022; 10.93 ± 0.024, 12.62 ± 0.023, 14.34 ± 0.025; 35.89 ± 0.049, 39.82 ± 0.040, 42.36 ± 0.043, 48.21 ± 0.037; 33.03 ± 0.033, 43.74 ± 0.06, 45.59 ± 0.033, 49.92 ± 0.029; 5.23 ± 0.015, 6.10 ± 0.016, 7.30 ± 0.015, 8.36 ± 0.015 and 89.89 ± 0.038, 101.54 ± 0.038 and 114.12 ± 0.037, respectively. Significant effects (P<0.01) of district and sex of bird on body weight and morphometric traits were observed. Age at first lay, annual egg production, clutch size and clutch interval were 323.02±0.302 days, 21.51±0.180 numbers, 9.87±0.098 numbers and 64.48±0.655 days, respectively. Two laying cycles, viz., September to October and December to February were observed. Egg weight, shell weight, shell thickness, shape index and specific gravity were 117.00±1.395 g, 14.19±0.225 g, 0.54±0.006 mm, 72.22±0.283 % and 1.09±0.001, respectively. Albumen Index, Haugh Unit, Yolk Index and yolk weight were 0.06±0.001, 59.79±0.726, 0.34±0.007 and 61.98±1.045, respectively. Age at slaughter (days), pre-slaughter weight (g), shrinkage (%), dressing percentage (%), evisceration loss (%) and ready to cook yield (%) in gander were 411.40 ± 4.26, 4192.80 ± 69.87, 4.36 ± 0.05, 67.10 ± 1.14, 7.58 ± 0.58 and 70.43 ± 1.05, respectively. The corresponding values in geese were 338.00 ± 21.48, 3878.40 ± 36.06, 4.76 ± 0.32, 66.11 ± 1.01, 7.02 ± 0.73 and 70.28 ± 1.24. In gander, yield of cut up parts on live weight (%) were 17.62 ± 0.19, 7.68 ± 0.14, 6.58 ± 0.10, 13.56 ± 1.29, 4.62 ± 0.12 and 8.05 ± 0.21 for breast, drumstick, thigh, back, neck and wings respectively. The corresponding values on dressed weight (%) were 27.49 ± 0.56, 11.98 ± 0.29, 10.26 ± 0.13, 21.01 ± 1.66, 7.21 ± 0.23 and 12.55 ± 0.25, respectively. In geese, cut up parts yield to live weight (%) were 17.58 ± 0.28, 6.74 ± 0.14, 6.71 ± 0.10, 12.40± 0.50, 4.35 ± 0.16 and 7.87 ± 0.33 for breast, drumstick, thigh, back, neck and wings, respectively. The corresponding values as per dressed weight (%) were 27.94 ± 0.42, 10.71 ± 0.19, 10.67 ± 0.16, 19.67 ± 0.43, 6.93 ± 0.33 and 12.48 ± 0.41, respectively. Meat to bone ratio of 2.05 ± 0.01 was found in both the sexes. 100 percent broody behavior observed. Fertility and hatchability (TES) were 91.38 ± 6.83 and 86.50 ± 7.59%. Mortality rate were 22.58, 15.24 and 11.72 at 0 to 1, 1 to 8 and 8 to 20 weeks. Screening for polymorphism in 5'-flanking region of PRL revealed transversion of G to C at location 117 and transition of C to T at location 182.The native geese attained much higher body weight at 6 months and 12 months of age compared to other poultry species. Various studies may be undertaken to genetically characterize the goose populations and association studies of genes with production traits may be conducted. Geese have promising role as alternate species for backyard poultry meat production. The detailed study would help in development of breed descriptors and baseline reference for future studies.ThesisItem Open Access STUDIES ON SWAMP BUFFALOES OF ASSAM UNDER FARM AND FIELD CONDITION(College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati, 2020-12) SARMA, MOMI; Das, G. C.The present investigation was conducted to study some of the morphometric, production & reproduction and milk constituent traits of swamp buffaloes of Assam under farm and field condition. Accordingly, the required data for farm animals were collected from the records maintained at the buffalo farm under Network project on swamp buffaloes of Assam, College of Veterinary Science, A.A.U. Khanapara and Government Livestock Farm, Department of A.H. & Vety., Govt. of Assam, Barhampur, Nagaon. The data for field animals were collected from different buffalo khutis located at Dhemaji, North Lakhimpur and Sivasagar districts of Assam through field survey by providing questionnaire as well as through personal interviews with the farmers. For morphometric traits, the measurements were made on adult buffaloes using standard measuring tape at farm as well as at field. For milk constituents milk samples of swamp buffaloes were collected randomly from farm as well as from field and were tested using Milk Analyzer, LAKTAN 1-4 (Model: 220). For morphometric traits 62 numbers of swamp buffaloes from Khanapara buffalo farm, 21 numbers from Barhampur buffalo farm and 224 numbers from field were considered. Out of 224 animals, 146 from Dhemaji and North Lakhimpur districts (north bank) and78 from Sivasagar district (south bank) were measured. For production and reproduction traits 1056 lactation records, of which 208 lactation records pertaining to 52 no. of animals from Khanapara buffalo farm and 848 lactation records pertaining to 212 no. of animals from field (buffalo khutis) covering all the three districts were considered. A total of 172 milk samples, out of which 46 samples from Khanapara farm and 126 from Dhemaji, North Lakhimpur and Sivasagar districts of Assam were tested for various milk constituents. The body measurement traits include body length, height at withers, heart girth and the pouch girth. The overall least-squares means (μ) for the body length, height at withers, heart girth and pouch girth were found to be 108.881 ± 1.448 cm, 126.793 ± 1.378 cm, 172.630 ± 2.608 cm and 174.760 ± 2.422 cm, respectively. Significant (P<0.05) effect of the two locations under study was observed in case of all the body measurement traits viz., body length, heights at withers, heart girth and pouch girth. Significant (P<0.05) effect of sex was seen for all the body measurement traits viz., body length, heights at withers, heart girth and pouch girth. Male buffaloes were observed to show higher values for all the traits of body measurements than the females. The head measurement traits include head length and eye to eye space. The overall least-squares means (μ) for the head length and eye to eye space was found to be 43.746 ± 0.772 cm and 15.125 ± 0.314 cm, respectively. Significant (P<0.05) effect of the two locations under study was observed in case of head length. Significant (P<0.05) effect of sex was seen on both the traits of head measurements viz., head length and eye to eye space. Male animals have shown higher values for both the traits of head measurements than the females. The least-squares means (μ) for the ear length was found to be 21.498 ± 0.287 cm. The horn measurement traits include circumference of horns at base, horn length, space between two horns at base and space between two horns at tips. The overall least-squares means (μ) for the circumference of horns at base and horn length was found to be 18.975 ± 0.491 cm and 43.312 ± 0.934 cm, respectively. While the overall least-squares means (μ) for the space between two horns at base and space between two horns at tips were found to be 16.293 ± 0.316 cm and 37.118 ± 0.807 cm. Significant (P<0.05) effect of sex was observed in case of horn length and space between two horns at base. Male buffaloes have shown higher values for the horn measurement traits. The tail measurement traits include length of tail and length of switch of tail. The overall least-squares means (μ) for the length of tail and length of switch of tail were found to be 69.956 ± 0.913 cm and 13.625 ± 0.298 cm. Significant (P<0.05) effect of sex was observed in case of length of tail. Male buffaloes have shown higher value for the length of tail than the female buffaloes. For colour characteristics, coat colour, horn colour and tail switch colour was included. Coat colour was predominantly grey (98.58%) with few white (albino) (1.52 %) was observed. For horn colour, black colour was observed in all the observations. For tail switch colour two variants were observed viz., black in 86.65 % observation and brown in 13.35% observation. The least-squares means (μ) for first lactation milk yield and for first lactation length was found to be 449.379 ± 5.517 kg and 239.443 ± 5.845 days, respectively. Highly significant effect (P˂ 0.01) of location on first lactation milk yield was observed. The least-squares means (μ) for first peak yield and for days to attain first peak yield was found to be 2.810 ± 0.016 kg and 59.204 ± 3.167 days, respectively. Significant influence (P˂ 0.05) of season of calving was observed on days to attain peak yield. The least-squares means (μ) for the age at first calving was found to be 53.714 ± 2.269 months. The least-squares means (μ) for first gestation length and first dry period were found to be 308.549 ± 1.444 days and 246.913 ± 6.452 days, respectively. Significant influence (P˂ 0.05) of sex of calf was observed on first gestation length. Highly significant influence (P˂ 0.01) of location was observed on first dry period. Significant influence (P˂ 0.05) of season of calving and sex of calf was observed on first dry period. The overall least-squares means (μ) for first service period and first calving interval were found to be 178.042 ± 5.852 days and 486.533 ± 6.879 days, respectively. Highly significant influence (P˂ 0.01) of location was observed on service period. Significant influence (P˂ 0.05) of season of calving and sex of calf was observed on first service period. Significant influence (P<0.05) of location was observed on first calving interval. The least-squares means (μ) for birth weight was found to be 29.441 ± 2.446 kg. The least-squares means (μ) for lactation milk yield and lactation length was found to be 453.654 ± 6.715 kg and 243.847 ± 6.394 days, respectively. Highly significant influence (P˂ 0.01) of location and order of lactation were observed on both lactation milk yield and lactation length. The overall least-squares means (μ) for peak yield was found to be 2.951 ± 0.035 kg and days to attain peak yield was found to be 59.626 ± 4.591 days. Highly significant influence (P˂ 0.01) of location was observed on days to attain peak yield. Highly significant influence (P˂ 0.01) of order of lactation was observed on peak yield as well as on days to attain peak yield. The least-squares means (μ) for gestation length and dry period was found to be 309.079 ± 1.387 days and 238.396 ± 5.664 days, respectively. Highly significant (P˂ 0.01) influence of sex of calf was observed on gestation length. Highly significant (P˂ 0.01) influence of order of lactation was seen on dry period. The overall least-squares means (μ) for service period and calving interval was found to be 177.709 ± 5.555 days and 486.700 ± 5.503 days, respectively. Highly significant (P˂ 0.01) influence of order of lactation was observed on calving interval. The overall least-squares means (μ) for milk fat %, solid-not fat %, milk protein % and total solids % were found to be 7.591 ± 1.148, 9.047 ± 0.052, 3.032 ± 1.126 and 16.638 ± 1.125 % respectively. Significant (P˂ 0.05) influence of location was observed on milk fat % and milk protein %.ThesisItem Open Access GENETIC STUDIES ON THE PERFORMANCE OF HD-K75 PIGS(College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati, 2022-02) BAYAN, JYOTISHREE; Zaman, Galib UzPig farming in India is primarily a small scale unorganized rural activity and is an integral part of diversified agriculture particularly in the tribal belt of the country and have been contributing to improve the livelihood of poor and socially weaker section of the society including the tribal people of India. Pig farming in Assam is rapidly gaining momentum in recent years as the farmers are finding it a profitable enterprise. The present investigation was carried out on HD-K75 pigs (75% Hampshire inheritance and 25 % indigenous inheritance) that are bred and maintained in the All India Coordinated Research Project (AICRP) on pigs, ICAR, located at College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati-781022. The data were collected and compiled from the progenies of 44 sires and 114 dams maintained over a period of 6 years from 2012 to 2018. The objective of the investigation were to study some important traits of growth and reproduction and the effect of various non-genetic factors influencing these traits, determine heritability, genetic correlation and phenotypic correlation of some important growth and reproduction traits, to construct linear, partial and multiple regression equations and to predict adult body weights based on early body weights. Traits included in the study were body weights at birth, weaning, 1st, 2nd, 3rd, 4th, 6thand 8th months of ages, and daily body weight gains during pre-weaning (birth to weaning) and post-weaning periods (from weaning to 4th, 6th and 8th month of ages), age at sexual maturity, gestation period, farrowing interval, litter size at birth, litter weight at birth, litter size at weaning and litter weight at weaning. The overall mean body weight in HD-K75 pigs at birth, 1 month, weaning, 2 months, 3 months, 4 months, 6 months and 8 months of age were 1.001 ± 0.001 kg, 6.912 ± 0.011 kg, 9.666 ± 0.013 kg, 12.207 ± 0.007 kg, 18.324 ± 0.001 kg, 28.349 ± 0.052 kg, 51.177 ± 0.091 kg and 71.229 ± 0.110 kg respectively. The average daily body weight gains (ADG) in g during birth to weaning, weaning to 4th month, weaning to 6th month and weaning to 8th month of ages were 206.299 ± 0.311, 237.294 ± 0.773, 299541 ± 0.700 and 311.206 ± 0.557 respectively. Least-squares analysis of variance revealed highly significant effect of season of birth, parity and sex on body weight at all ages and daily body weight gains at various stages of growth. Piglets born during winter season had higher body weight at birth, 1 month, 42 days, whereas piglets born during post-monsoon season had higher body weight at 2 months, 3 months, 4 months and 6 months of age and piglets born during monsoon season had higher body weight at 8 months of age. Pre-weaning ADG was higher in piglets born during monsoon season and lowest in animals born during pre-monsoon season. The ADG during 42 days to 4th month and 42 days to 6th month were found to be higher in piglets born during post-monsoon season and lowest in piglets born during pre-monsoon season. Piglets born in second parity had higher body weight at 3 months, 4 months, 6 months and 8 months whereas piglets born in third parity had higher body weight at birth and 1 month and piglets born in first parity had higher body weight at weaning and 2 months of age. Significantly higher ADG was found during pre-weaning period of growth in animals born in first parity. Further, the ADG from 42 days to 4th month, 42 days to 6th month and 42 days to 8th months of age were found to be higher in animals born in second parity and revealed significant difference with the animals of first and third parities. Males showed higher body weight and higher ADG than females in all stages of growth. ii The h2 estimates for body weight and ADG at various stages of growth in the present study were moderate to high in most of the cases which indicated that the population under study possess good amount of additive genetic variance and there is scope of genetic improvement of the herd in terms of growth traits and ADG at various stages using adequate methods of selection by incorporating these estimates. The phenotypic correlations among body weights at various ages were moderate and positive in some cases and negative in some cases. The phenotypic correlations among ADG at various stages of growth were moderate and positive in some cases and negative in some cases. The genetic correlations among body weights at various ages of growth were low to high and positive in some cases and negative in some cases. The genetic correlations among ADG at various stages of growth were moderate and positive in some cases and negative in some cases. The overall means for age at sexual maturity, gestation length and farrowing interval were found to be 205.294 ± 1.054, 112.70 ± 0.119 and 216.781 ± 2.565 days respectively. Effect of season of birth on age at sexual maturity was found to be non-significant as indicated by least square analysis of variance. Non-significant effect of season of birth and parity on gestation period was observed in the present study in HD-K75 pigs. Least-squares analysis of variance revealed significant difference between the various seasons under study in respect of farrowing interval. Sows that farrowed during post-monsoon season had significantly longer farrowing interval and differed significantly with other seasons and the shortest farrowing interval was observed in sows that farrowed during monsoon season. Least square analysis of variance revealed that the effect of parity on farrowing interval in the present study was non-significant. The h2 estimates of age at sexual maturity, gestation length and farrowing interval for HD-K75 pigs was low to moderate in magnitude. The phenotypic correlation among the reproduction traits were found to be low and positive in most cases. The genetic correlation among the reproduction traits were found to be moderate and positive in most cases. The overall means for litter size at birth, litter size at weaning, litter weight at birth and litter weight at weaning were found to be 7.747 ± 0.088, 7.556 ± 0.093 numbers and 7.804 ± 0.084 and 74.644 ± 0.0886 kg respectively. Least-squares analysis of variance revealed significant effect of season of birth on litter size at birth, litter size at weaning, litter weight at birth and litter weight at weaning. Piglets born during pre-monsoon season showed highest litter size at birth and weaning. Piglets born during post-monsoon season and winter season showed highest litter weight at birth and litter weight at weaning. Further, litter size at birth, litter size at weaning, litter weight at birth and litter weight at weaning were found lowest during monsoon season. Least-squares analysis of variance revealed that parity had significant effect on litter size at birth, litter size at weaning, litter weight at birth and litter weight at weaning. Piglets born in third parity showed highest and piglets born in first parity showed lowest litter size at birth, litter size at weaning, litter weight at birth and litter weight at weaning respectively. The h2 estimates of litter size at birth and litter size at weaning in HD-K75 pigs in the present study were found to be low in magnitude. The h2 estimates of litter weight at birth and litter weight at weaning were found to be medium in magnitude. The phenotypic correlations among the litter traits were found to be high and positive in most cases. The genetic correlations among the litter traits were found to be moderate to high and positive in most cases. Prediction equations were developed using post-weaning body weights at 6 months (Y1) and 8 months (Y2) of ages as dependent variables based on pre-weaning body weights at birth (X1), 1 month (X2), 42 days (X3), 2 months (X4) and 3 months (X5) of age as independent variables in various combinations. For prediction of adult body weights on the basis of body weights at early ages in HD-K75 pigs, linear, partial and multiple regression equations were developed. The linear regression equations for predicting body weights at 6 months of age had comparatively higher R2 values than 8 months of age in HD-K75 pigs, but were not found to be reliable as the R2 values were less than 60 %. The partial regression equations for predicting body weights at 8 months of age had comparatively higher R2 values than 6 months of age and were found to be more reliable. All the multiple regression equations developed to predict the adult body weight at 8 months of age can be reliably used when 3-5 independent variables are considered. Multiple regression equations developed to predict 8 month body weight showed highest reliability with R2 values ranging from 70.87 to 90.56 %. High R2 values indicates that the 8 month body weight can be predicted more efficiently than 6 month body weight. The multiple regression equations for predicting body weight at 8 months using 3 independent variables viz. body weight at birth, 1 month and weaning showed high R2 value of 90.21 % which was found to be highly reliable and the best combination, though higher R2 values were obtained in multiple regression equations using 4 and 5 independent variables.
