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National Dairy Research Institute, Karnal
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ThesisItem Open Access PERFORMANCE AND BEHAVIOUR OF FRESHLY CALVED BUFFALOES SUBJECTED TO STAGGERED INTEGRATION IN PAIRS INTO A GROUP OF LOOSE HOUSED LACTATING BUFFALOES(ICAR-NDRI, KARNAL, 2024) SUNITA DIDEL; M. L. KAMBOJBuffaloes, being highly social animals, often face stress during regrouping due to changes in social hierarchy and agonistic interactions. To address this, the present investigation was carried out with the objectives of studying the effects of staggered entry of freshly calved buffaloes in pairs into an established group on their behaviour, social stability, productive performance and milk quality. For this study, a lactating herd of 70-80 Murrah buffaloes maintained together in a loose house was considered as a resident group of animals. In this resident group, 36 freshly buffaloes allotted at random to two groups were introduced 5 days post-partum spread over 6 months (from June, 23 to October, 2023) in order of their calving. The buffaloes in one group (n = 12) were directly introduced individually into the resident group of buffaloes as per existing farm practice (control, T0). The buffaloes in treatment (T1) group (n = 24 in 12 pairs) were introduced into the resident group in pairs in a staggered manner over 4 days. On day 1, 2 and 3 these buffaloes were kept in the resident group for 2, 4 and 8 hours respectively and were withdrawn afterwards and housed separately where they were provided similar conditions of feeding and housing for the rest of the time. On day 4, the buffaloes were fully integrated into the resident group. The data on the mean differences in behavioral, social stress, performance and milk quality parameters of the two groups of buffaloes recorded over 100 days post-partum were analysed by performing one-way analysis of variance in SSPS statistics 20. Results revealed that T1 buffaloes exhibited higher (P<0.05) daily feeding time (340.81Ā±6.55 min/d) and rumination time (432.52Ā±9.53min/d) compared to T0 (319.56Ā±9.38; 390.66Ā±15.04 min/d) respectively. Water drinking time was significantly greater (P<0.01) in T1 (13.46Ā±0.57 min/d) than T0 (10.79Ā±1.00 min/d). T1 also displayed increased (p<0.05) total lying time (611.09Ā±6.11 min/day) compared to T0 (558.50Ā±10.86 min/d). Conversely, T0 exhibited higher (P<0.05) standing time (881.50Ā±10.86 min/d) and idle standing time (355Ā±23.53 min/day) than T1 (828.93Ā±6.10; 283.22Ā±25.69 min/d) respectively. Agonistic interactions were significantly higher in T0 as compared to T1. Notably, there were no differences in the mean daily times spent on feeding, rumination, resting and number of agonistic interactions in the two groups of buffaloes after 3-4 weeks of introduction into the resident group indicating their social integration into the resident group. Additionally, T1 buffaloes demonstrated higher (p<0.05) daily milk yield (8.91 Ā±0.67 kg/d) and milk flow rate (0.977Ā± 0.04 kg/min) compared to T0 (6.17Ā±0.98 kg/day, 0.861Ā±0.037 kg/min). Milk quality also favoured T1, showing lower (p<0.05) somatic cell counts (2.01Ā±0.11 lakhs/ml) and higher fat percentage (6.82Ā±0.17%) compared to T0 (2.58Ā±0.13 lakhs/ml, 5.98Ā±0.43%) respectively. Furthermore, plasma cortisol levels were significantly (p<0.05) lower in T1 (6.59Ā±0.59 ng/ml), indicating reduced stress compared to T0 (12.21Ā±0.80 ng/ml). In conclusion, staggered entry of freshly calved buffaloes in pairs positively influenced behaviour, social interactions, and productive performance. Gradual introduction in pairs resulted in improved behaviour, enhanced milk yield, better milk quality, and reduced stress compared to direct single integration into the resident herd.ThesisItem Open Access EVALUATION OF METHANE MITIGATION POTENTIAL OF HERBAL FEED ADDITIVES UNDER IN VITRO MODEL(ICAR-NDRI, KARNAL, 2024) SIMRAN KAMBOJ; GOUTAM MONDALHerbaceous plants have garnered substantial attention for their potential role in mitigating methane emissions in ruminants, given their complex phytochemical composition. The current study investigates the methane-mitigating potential and impact on rumen fermentation parameters of seven herbaceous feed additives, namely Andrographis paniculata, Swertia chirata, Artemisia annua, Mangifera indica seed kernel, Carica papaya leaves, Euphorbia thymifolia, and Heliotropium indicum. Additionally, a synergistic herbal formulation comprising Andrographis paniculata, Mangifera indica seed kernel, and Carica papaya leaves have been evaluated at 6 different inclusion levels- 0%(control), 1%, 2%, 5%, 7%, 10% in 3 TMRs with NDF levels 30, 40, 50%. The investigation unfolds in two phases: an initial screening of individual additives followed by an assessment of the composite herbal blend. Andrographis paniculata manifests a noteworthy reduction in methane production, showcasing a decrease of 17% at 10% level of inclusion, at NDF 30. Concurrently, total gas production and in vitro dry matter digestibility (IVDMD) register significant declines, underscoring its positive impact on rumen fermentation. Swertia chirata exhibits methane reduction of 8% at 10% inclusion, 6% at 7% inclusion, and 4% at 5% inclusion in NDF 30. Alterations in total gas production and short-chain fatty acid (SCFA) profiles imply a potential role in methane mitigation. Artemisia annua consistently diminishes methane production across all inclusion levels, with reductions of 12% at 10%, 9% at 7%, and 6% at 5% in NDF 30. Progressive increases in total gas production underscore its promise for methane mitigation. Mangifera indica seed kernel demonstrates a reduction in methane production of 19% at 10%, 13.5% at 7%, and 10% at 5% in NDF 30. Observable changes in total gas production and SCFAs, particularly propionate, highlight its impact. Carica papaya leaves reveal a substantial reduction in methane production of 21% at 10%, 13% at 7%, and 9% at 5% in NDF 30. Alterations in total gas production and SCFAs suggest a discernible influence on fermentation patterns. Euphorbia thymifolia exhibits a methane reduction of 3-5% at 5-10% inclusion in NDF 30, with minimal changes in NDF 40 and NDF 50. Heliotropium indicum demonstrates a methane reduction of 3-4% at 5-10% inclusion in NDF 30, with minimal changes in NDF 40 and NDF 50. The herbal formulation, comprising Andrographis paniculata, Mangifera indica seed kernel, and Carica papaya leaves, manifests a substantial reduction in methane of 23% at 10%, 15% at 7%, and 9% at 5% in NDF 30. Negligible changes are observed at NDF 40 and 50, indicating effective methane mitigation without compromising digestibility. In conclusion, both individual herbal plants and the admixture of the plants are effective in methane mitigation at 5-10% inclusion levels, thereby positioning them as promising eco-friendly alternatives in ruminant nutrition.ThesisItem Open Access EFFECT OF DIETARY CURCUMIN SUPPLEMENTATION ON SEMINAL ATTRIBUTES OF CROSSBRED BULLS DURING INDUCED HEAT STRESS(ICAR-NDRI, KARNAL, 2023) MANISHA YADAV; NISHANT KUMARAn imbalance between heat production in the body and its dissipation is called as state of heat stress. When environmental temperature is high, it challenges the animalās capacity to maintain energy, thermal, water, hormonal and mineral balance ultimately resulting in heat stress. Since bull fertility is considered to be adversely affected by hot humid condition, the current increase in global temperature as a result of climate change may have profound implications for future livestock production. Any strategy which could reduce the adverse effect of heat stress on bull fertility would be a welcome innovation. The present study was conducted with objectives (i) to study the effect of dietary curcumin supplementation on seminal and biochemical attributes in crossbred bulls and (ii) to evaluate the effect of dietary curcumin supplementation on physiological and sexual behavior of crossbred bulls. Heat stress was induced by placing the animals in psychrometric chamber. Both control and supplemented group were exposed to a temperature and relative humidity 37Ā±20C and 50% respectively for 8 hours (9 AM to 5 PM) per day for 15 days. Pre and post heat exposure semen samples were evaluated during the experiment. The results indicated that the ejaculate volume was not significantly altered by induced heat stress. Sperm concentration, mass activity, motility, viability, plasma membrane integrity and sperm morphology at fresh and post thaw stage were significantly (P<0.05) altered by heat stress. However, these adverse effects were significantly (P<0.05) reduced in Curcumin supplemented groups. There was also a significant (P<0.05) difference in respiratory rate, rectal temperature and skin surface temperature between control and supplemented groups. There were significant (P<0.05) difference in the serum cortisol and testosterone concentration in both groups. Heat stress deteriorates semen quality from 2nd week and recovery occurs after 5th-6th week in supplementedgroup and after 8th week in control group and alters some of the physiological parameters. It can be concluded that supplementation with Curcumin have been found to ameliorate adverse effects of heat stress in cross bred bulls. The nutritional supplements like curcumin to ameliorate the adverse effects of heat stress in animals could be a beneficial and novel strategy particularly for animals during heat stress.ThesisItem Open Access EFFECT OF MANGO SEED KERNEL AND RAMIE FODDER ON IN-VITRO RUMEN FERMENTATION, METHANE PRODUCTION AND RUMEN MICROBIAL PROFILE(ICAR-NDRI, KARNAL, 2023) MANISHA; ANJALI AGGARWALIt is essential to investigate novel approaches to decrease greenhouse gas emissions from ruminants in order to support livestock production and mitigate global warming. This study was conducted in 3 phase in-vitro trials to examine the effect of Ramie fodder (replaced by berseem at 10%, 20%, 30%, 40%, 50 % level), Mango Seed Kernel (1%, 3%, 5%, 7% and 10% level) and combination of best two levels of Ramie fodder with various levels of MSK under in-vitro condition. Total mix ration (TMR) with 60% roughage and 40% concentrate mixture was formulated as control group. In phase 1st results revealed that total gas production, methane production, In-vitro dry matter digestibility (IVDMD) and Invitro organic matter digestibility (IVOMD) decreased continuously with increased level of replacement, as compare to control group. However, a maximum reduction 12.7% and 14.95% in methane was seen at 40% and 50% replacement respectively. Based on the overall results of in-vitro studies, 40% and 50% inclusion level of Ramie fodder was selected for further combinations studies. In 2nd phase on addition of mango seed kernel (MSK) in TMR, total gas production and methane production decreased with increased level of MSK. The mean value of IVDMD and IVOMD reduced significantly (p <0.05) at 10% level. Phase 3rd trial was conducted to study the combination effect of Ramie fodder with MSK (40% Ramie+1/3/5/7/10% MSK and 50% Ramie+1/3/5/7/10% MSK) the results revealed that 40% Ramie+various levels of MSK, induced significant (P<0.05) reduction in methane production in all treatment groups as compare to control group. However, the maximum methane reduction (27.64%) was seen at 40% Ramie+10% MSK. The combination of 50% Ramie+various levels of MSK, reduced total gas and methane production in all treatment groups, and maximum methane reduction (36.74%) was seen at 40% Ramie+10% MSK. Total gas production reduced significantly (P<0.05) at 3%, 5%, 7% and 10% MSK+ 40%/50% Ramie. IVDMD reduced significantly at 7% and 10% MSK + 40%/50% Ramie whereas, in IVOMD significant (P<0.05) reduction was seen at 10% MSK + 40%/50% Ramie. The activity of carboxymethyl cellulase and acetly esterase reduced significantly (p <0.05) at 7% and 10% MSK+40%/50% Ramie. The population of methanogens reduced significantly (p <0.05) for all treatment groups of combination compared to control group, whereas the population of total fibrolytic bacteria reduced significantly (p < 0.05) at 7% and 10% MSK+50% Ramie. Protozoal population reduced significantly (p < 0.05) at 3% , 5% and 7% MSK+50% Ramie, as compared to control group. Therefore, it can be concluded that the of 5% MSK in combination with 50% Ramie fodder offers a highly effective strategy for reducing enteric methane emissions by 35.99%, while maintaining key rumen physiological parameters under in-vitro condition.ThesisItem Open Access IMPACT OF CUMIN AND PRILL FAT SUPPLEMENTATION ON GROWTH PERFORMANCE AND STRESS MARKERS OF CROSSBRED FEMALE KIDS DURING DIFFERENT SEASONS(ICAR-NDRI, KARNAL, 2023) ROOHANI SHARMA; SOHAN VIR SINGHThe experiment was conducted to study the impact of cumin and prill fat supplementation on growth performance and stress markers of crossbred (Alpine x Beetal) female kids during different seasons. Twenty-four crossbred kids (3 months) were selected from Livestock Research Centre, ICAR-National Dairy Research Institute, Karnal (Haryana) and divided equally into four groups having six kids in each group i.e., T1 (controlā basal diet), T2 (basal diet+ cumin @ 2% of conc. fed), T3 (basal diet+ prill fat @ 2.5% of DM intake) and T4 (basal diet+ combination of cumin @ 2% of conc. fed and prill fat @ 2.5% of DM intake). The environmental variables viz- dry and wet bulb temperature and relative humidity were recorded and temperature humidity index (THI) was calculated. Physiological parameters and infrared thermography images of all the group of kids were recorded at weekly interval. Blood sampling, DMI, environmental variables and growth measurements were recorded at fortnight interval. The rectal temperature and respiration rate of crossbred goat kids was significantly (P<0.05) higher in control group as compared to the treatment groups during different seasons. The overall mean values of RR and RT of all groups was highest during hot humid season, followed by hot dry and spring season. The control group's ear pinna and dorsal surface temperature was significantly (P<0.05) higher as compared to treatment groups during different seasons. The blood Hb and PCV exhibited significantly (P<0.05) higher levels in the combination and prill fat supplemented groups compared to the control group during different seasons. No significant (P< 0.05) difference was observed in the overall mean values of plasma albumin, globulin, total protein, ALT and AST during different seasons among all the groups. The overall mean values of plasma glucose level showed significantly (P<0.05) higher values in treatment groups as compared to the control group during different seasons. The plasma cortisol hormone was significantly (P< 0.05) lower whereas, overall mean values of TSH and growth hormone was significantly (P< 0.05) higher in treatment groups as compared to control. TSH and cortisol showed significantly lower and higher levels during summer season respectively compared to spring season. Morphological parameters viz. heart girth, body length and height at wither showed significantly (P<0.05) higher values in prill fat and combination supplemented group as compared to cumin and control group during different seasons. The overall mean values of absolute weight were significantly (P<0.05) lower in control compared to treatment groups during different seasons. The ADG of treatment groups were significantly (P<0.05) higher than control group during different seasons. ADG was found to be significantly (P<0.05) lower throughout the summer season (hot dry and hot humid) compared to the spring season. DMI was significantly (P<0.05) higher in the cumin and combination supplemented groups compared to the control group, with the lowest DMI found during the hot humid season. The supplementation of cumin, prill fat and their combination helped in lowering stress markers (cortisol, RT, RR), increasing average daily gain, dry matter intake, growth hormone and TSH, and ultimately improved growth performance of kids of crossbred goat. The body weight gain was found to be significantly (P<0.05) higher in combination supplemented group however, this supplementation was less profitable compared to prill fat and cumin supplemented group since the input cost was higher. Therefore, prill fat supplementation can be recommended to the goat keepers for better returns.ThesisItem Open Access ESTABLISHMENT of BETA-LACTOGLOBULIN KNOCKOUT BUFFALO CELL LINES using CRISPR-RNP COMPLEXES and INDIGENOUS LAB-MADE TRANSFECTION BUFFER(ICAR-NDRI, KARNAL, 2023) TARA ASEEM PUSHPA; N. L. SELOKARBeta-Lactoglobulin (BLG) is a major milk allergen present in farm animals; but it is absent in human milk. Engineered endonucleases, including CRISPR, enable targeted genetic modification in livestock. In India, the importance of buffaloes in the dairy industry is pronounced, accounting for approximately 45% of the country's milk production. This highlights the dominant position of buffaloes in fulfilling the milk requirements of the Indian population. The production of -lactoglobulin knockout buffaloes could have significant implications, including the availability of hypoallergenic milk and increased demand for buffalo milk among individuals with milk allergies. By considering this in mind, the present thesis study was designed to establish -lactoglobulin knockout cell clones, and the production of -lactoglobulin-gene edited embryos. To achieve targets of this study, five major experiments were performed: 1) Designing of CRISPR guides against BLG gene; 2) Determination of editing efficiencies of designed guides using different methods such as T7E assay, TIDE, and ICE analysis; 3) Generation of single cell clones from electroporated cells; 4) Validation of mono-allelic or bi-allelic mutations in single-cell colonies; 5) Production of cloned blastocyst stage embryos from edited BLG-gene edited cells and assessment of their quality. In summary, a total three sgRNAs were designed, of which two sgRNAs were efficient to edit the BLG locus with indigenous lab-made buffer. With the best optimized condition, in the present study, the seven BLG-gene edited single cell clones were established that are 1) C1 has bi-allelic heterozygous mutation, one allele with -107 bp deletion and the other allele with - 9bp deletion with C/T conversion; 2) C2 has bi-allelic homozygous mutation, both alleles have -32 bp deletion; 3) C4 has mono-allelic mutation, in which one allele has -151 bp deletion; 4) C8 has the mono-allelic mutation, in which one allele is with -8 bp deletion; 5) C10 has bi-allelic heterozygous mutation, one allele with -150 bp deletion and the other allele with one bp T insertion; 6) C11 has mono-allelic mutation, in which one allele is with -14 bp deletion; and 7) C13 has bi-allelic heterozygous mutation, one allele with -27 bp deletion and the other allele with - 26 bp deletion. Following the use of some edited single cells clones as nuclear donor for SCNT applications, the embryonic developmental rates were different among the cell type; however, the total cell number and apoptotic index were similar among edited and non-edited (wild type) donor cells. Thus, SCNT outcome indicated that genetic modification using CRISPR did not affect the cloned embryonic developmental competence. The results of this study demonstrated the successful establishment of a pipeline for CRISPR-based manipulation of the -lactoglobulin locus in buffaloes and the production of BLG-gene-edited cloned embryos. The generated -lactoglobulin knockout cell clones will provide a valuable resource for the production of live genome-edited buffaloes.ThesisItem Open Access PREDICTION OF THERMAL STRESS IN INDIGENOUS AND CROSSBRED CATTLE USING INFRA-RED THERMOGRAPHY AND EMERGING MACHINE LEARNING TECHNIQUES(ICAR-NDRI, KARNAL, 2023) AFREED MUHAMMED, N. V.; SOHAN VEER SINGHThe present study was conducted at the Livestock Research Centre (LRC) of ICAR-NDRI, Karnal to study the thermogram and physiological responses of Karan Fries and Tharparkar cattle during different seasons and to develop and validate machine learning models for intelligent prediction of thermal stress (rectal temperature) in the Karan Fries and Tharparkar cattle. Twenty each of Tharparkar and Karan Fries cattle were selected and managed under standard managemental practices followed at LRC and fed as per the ICAR standard (2013). The research work was carried out during the extremes of winter (December to January), spring (February to March) and summer (May to June) season. The physiological parameters and infrared thermography of experimental animals was recorded twice weekly and the blood samples were collected fortnightly from six animals of each breed for the haematological and biochemical parameters. The environmental parameters viz dry and wet bulb temperature, black globe temperature, wind velocity, relative humidity and temperature humidity index and black globe humidity index was calculated from the recorded environmental parameters. Infrared thermograms were analyzed for the skin temperature at different anatomical regions i.e. muzzle, forehead, eye, ear base, ear tip, bridge of nose, neck, brisket, hump, dorsal, ventral, fetlock, knee, elbow, shoulder and lateral body. The environmental, physiological and infrared thermography readings were used for the development of the predictive models by different machine learning algorithms such as Bayesian Regularized Neural Network (BRNN), K-Nearest Neighbors (KNN), Support Vector Machine (SVM) and Multiple Linear Regression (MLR).There was significant (P>0.05) difference in mean values of physiological parameters viz RT and RR being higher during summer followed by spring and winter season within the breeds. RT was significantly (P<0.05) higher in Karan Fries than Tharparkar cattle during different seasons. Whereas, RR was significantly (P<0.05) higher in Karan Fries than Tharparkar cattle during spring and summer season. Within the breed a significant (P<0.05) difference was observed in mean values of thermal imaging temperature at different anatomical sites viz. muzzle, forehead, eye, base of ear, tip of ear, bridge of nose, neck, brisket, hump, dorsal, ventral, fetlock, knee, elbow, shoulder and lateral body region during different season. These values of skin temperature were highest during summer followed by spring and winter season. Among the both breeds of cattle, thermal imaging temperature at different anatomical sites viz. brisket, ventral and shoulder regions was significantly (P<0.05) higher in KF than TP during winter season. During the spring season a significant (P<0.05) difference was observed in the mean values of the muzzle and neck region temperature among the breeds. Significantly (P<0.05) lower mean values of red blood cell (million/mm3) and haemoglobin content (gm %) was found in both breeds during summer season compared to the other seasons. Within breed and among the breeds, a significant (P<0.05) difference was observed in mean values of plasma cortisol levels (ng/ml) during different seasons. Within the breed of cattle, a significant (P<0.05) difference was found in mean values of catalase activity (Ī¼mole of H2O2 consumed/min/mg Hb) during different seasons and values also differed significantly (P<0.05) among Karan Fries and Tharparkar cattle during spring and summer season. There was a (P<0.05) significant difference in mean values of superoxide dismutase activity (u/mg/Hb) during different seasons within the breeds. The levels of both the antioxidant enzymes were higher in KF than TP during all the seasons. The mean values of malondialdehyde (MDA) (nmol/mg) differed significant (P<0.05) within the breeds during different seasons and also differed significantly (P<0.05) among the both breeds of cattle during winter seasons. The prediction models developed by using Bayesian Regularized Neural Network (BRNN), K-Nearest Neighbors (KNN), Support Vector Machine (SVM), and Multiple Linear Regression (MLR) algorithms. The animal parameters like breed, infrared temperature of eyes, the respiration rate and the environmental stress indicator temperature humidity index (THI) used as input variables for the prediction of the rectal temperature/ heat stress. The four input parameters were selected based on the correlation among the parameters to the rectal temperature. The final predictive model by KNN algorithm exhibited the mean absolute error of 0.1627, root mean square error of 0.2059 and the R2 value of 0.7788 (R value as 0.8825), which is comparatively higher than the other models developed in this study. The BRNN model got the R2 value of 0.75 and the SVM and MLR models got R2 of 0.74. both the classical and machine learning models works well, but the machine learning model (KNN , BRNN, SVM) works slightly better than the classical Multiple Linear Regression models. Based on the results obtained, it can be concluded that the physiological parameters (thermal imaging temperature of eye and respiration rate) and THI are the most accurate parameters for the prediction of stress levels (Rectal temperature) of cattle.ThesisItem Open Access FLOW CYTOMETRIC ANALYSIS OF MILK FOR IDENTIFICATION OF UDDER HEALTH DURING EARLY LACTATION IN HIGH AND LOW PRODUCING COWS(ICAR-NDRI, KARNAL, 2023) ANISH KOUL; A.K. DANGMastitis, an inflammatory condition of the mammary gland and is classified as clinical or subclinical depending on the visibility of the effects of inflammation of the mammary gland. The present study was undertaken to study the total and differential milk cell counts and udder surface temperature in subclinical and clinical mastitis in both indigenous and cross bred cows. A total of 72 cows were selected from Livestock Research Center (LRC), ICAR- NDRI, Karnal and divided into two groups, 36 indigenous (Sahiwal) and 36 crossbred (Karan Fries) cows, and further subdivided into three groups viz., healthy (n=12), subclinical (n=12) and clinical mastitis (n=12) according to their somatic cell count (SCC) and California mastitis test (CMT). Milk from each cow was collected hygienically and evaluated for milk composition, SCC, and DLC. DLC was done by two methods, first by manual microscopic method and then by flowcytometry method by using specific antibodies. Macrophage and lymphocyte percentages decreased with increasing milk SCC, while neutrophil percentages increased during subclinical and clinical mastitis in both the breeds when DLC was done by manual microscopic method. Similar results were observed for neutrophils and macrophages percentage when it was done using flow cytometric method. However, for lymphocytes, two cells i.e., CD4 T helper cells and CD8 cytotoxic cells were evaluated by flowcytometric method. CD4 T helper cells and CD8 cytotoxic cells increased significantly from healthy to sub clinical cases and they decreased from sub clinical to clinical cases in both group of animals but not significantly. Also, ratio of neutrophil to macrophage (N:M) helps in better understanding of mastitis. Udder health was also found to be correlated with udder surface temperature as per objective of this study. It was observed that udder surface temperature increased significantly from healthy to sub clinical to clinical cases in both the group of the animals. It was also observed that with increasing SCC i.e., with increasing udder infection, udder surface temperature increased. This study indicated that DLC by flow cytometric technique and udder surface temperature can give a vivid picture of the condition of udder health.ThesisItem Open Access EVALUATION OF THE IMPACT OF MITOCHONDRIAL DNA D LOOP CHANGES ON MILK PRODUCTION TRAITS OF MURRAH BUFFALO(ICAR-NDRI, KARNAL, 2023) AMPADI, A.N; SADEESH, E.M.This study aimed to investigate the utility of mtDNA D-loop variations as predictive indicators of milk production variability in Murrah buffalo, a renowned breed known for its high milk yield, rich in fat and protein content. Lactation, a biologically energy-demanding process, primarily relies on mtDNA. The bovine mtDNA is approx16.3 kb extra-chromosomal circular genome, that serves as the primary energy source within the cell. The D-loop is a segment of mtDNA devoid of functional gene transcripts and plays a pivotal role in replication and transcription control while exhibiting susceptibility to mutations. In this study, the milk samples were collected from 50 Murrah buffaloes during a 30-day period. Physiological parameters, including milk yield, fat percentage, and protein percentage, were measured to calculate Energy Corrected Milk (ECM) for each animal. Based on the ECM data the animals were classified into three distinct groups: Group 1 (low ECM), Group 2 (medium ECM), and Group 3 (high ECM). Somatic cells were isolated from representative milk samples within each group, and total DNA extraction was performed from the somatic cells. Specific primers were employed to amplify the mtDNA D-loop, followed by sequencing. Sequences obtained were analyzed individually for each ECM group to identify potential variations. Forty-one sequences, each spanning 827 base pairs, were analyzed, with 11 sequences from Group 1, 25 from Group 2, and 5 from Group 3. Notably, Group 2 exhibited a higher prevalence of SNPs, while Group 3 displayed an increased occurrence of insertions and deletions (INDELs). Group 1 demonstrated the highest frequency of transitions, whereas Group 3 had the greatest percentage of transversions. A consistent deletion of āCā at the 714th position was observed in both Group 1 and Group 3, with a 68% prevalence in Group 2. Furthermore, unique variations specific to each group, as evidenced by negative Tajima D values, were identified, with Group 1 presenting the highest number of unique variations. While specific SNP associated with Group 2 was noticed. Nucleotide variations within the D-loop region may exert a notable impact on the mtDNA replication process, consequently leading to variations in mtDNA copy numbers among distinct animals. These fluctuating mtDNA copy numbers serve as indicative markers of cellular energy efficiency within tissues. Therefore, a comprehensive investigation aimed at assessing the influence of diverse cytoplasmic backgrounds in lactating Murrah buffalo on mtDNA copy number is essential. Additionally, an exploration of mtDNA isomorphism within coding regions holds promise for facilitating mt DNA-based selection strategies in breeding programs. Such strategies hold the potential to enhance milk production and optimize genetic traits associated with the production of this valuable breed.
