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ThesisItem Restricted DEVELOPMENT OF RECOMBINANT FLAGELLAR PROTEIN BASED IMMUNODIAGNOSTIC ASSAY FOR DETECTION OF Campylobacter jejuni(2021) BOBADE SUMEDHA SURESH; TANUVAS; VIJAYARANI K; TIRUMURUGAAN KG; THANGAVELU A; VAIRAMUTHU SCampylobacter spp. is a zoonotic pathogen and most reported cause of human gastroenteritis worldwide. Campylobacter jejuni is a commensal organism of the intestinal tract of poultry and other agriculturally important animals. The major transmission routes of Campylobacteriosis in human are consumption of contaminated or undercooked meat, raw milk or improper pasteurized milk. This study was undertaken to investigate the incidence of Campylobacter jejuni in milk, meat and faecal samples. A total of 143 samples comprising mastitis milk (20), raw milk (52), faecal swab of dog (19) suffer from diarrhoea, raw chicken meat (19), meat sample from sheep (5) and goat (3), beef sample (25) were collected aseptically. The samples were studied for cultural, biochemical and molecular examinations. Out of 143 samples 123 samples showed growth on modified blood free charcoal cefoperazone deoxycholate (mCCDA) agar media and 75 (60.97 %) isolates were identified as Campylobacter based on their morphology, Gram`s staining and motility. The isolated bacteria were characterized as C. jejuni based on biochemical tests viz. oxidase test, catalase test, nitrate reduction test, triple sugar iron test (TSI), hippurate hydrolysis and ninhydrin test specific for identification of C. jejuni and 47 (38.21 %) isolates were confirmed as C. jejuni. Based on triple sugar iron test 31 isolates were negative and catagorised as C. jejuni biotype 1 while 16 isolates were positive as biotype 2.ThesisItem Restricted A PROTEOMIC APPROACH TO AUGMENT QUALITY OF CHICKEN MEAT THROUGH DIETARY NATURAL ANTIOXIDANTS(2021) DIVYA MANJARI K; TANUVAS; PARTHIBAN M; KARUNAKARAN R; APPA RAO V; SENTHILKUMAR TMAProbiotics were reported to improve body weight gain and feed efficiency and reduced mortality in broiler chicken. In continuation of using probiotics as performance enhancers in broilers prebiotics are also supplemented to enhance the activity of probiotics and also aid in scavenging free radicals that are responsible for creating stress in the mitochondria. Here in this study chicory is a natural antioxidant that is used as prebiotic as feed supplement. In this study a total of 21 probiotic organisms belonging to seven different species were isolated from chicken gut. These twenty one probiotic organisms were characterized by Gram staining, Biochemical methods using 21 sugars, pH tolerance and bile salt tolerance test, enzyme analysis using 19 different enzymes. These probiotic organisms were identified using molecular studies using PCR amplification of 16S -23S spacer region and sequencing analysis. Based on the sequencing data, seven different species were found to exist namely Enterococcus faecalis, Enterococcus faecium, Enterococcus hirae, Enterococcus durans, Lactobacillus salivarius. Bacillus subtillis and Pediococcus acidilactici. Based on extracellular and in vitro assays a scoring system was evolved to evaluate the potential probiotic bacteria with antioxidant property. These probiotic bacteria along with prebiotic chicory was supplemented in broilers and the performance was studied and observed that the live weight was increased by 200 grams in the synbiotic treated groups. The pH, meat tenderness and total protein were also improved in synbiotic treated groups. Also the proteome of the breast muscle was studied for differentially expressed proteins which relate to the enhancement of broiler performance in the experimental trial.ThesisItem Restricted MOLECULAR CHARACTERIZATION OF MYCOBACTERIUM TUBERCULOSIS COMPLEX STRAINS FROM RUMINANTS AND EVALUATION OF ITS PURIFIED PROTEIN DERIVATIVE(2022) JAWAHAR A; TANUVAS; VIJAYARANI K; SARATHCHANDRA G; PAZHANIVEL N; MANOHARAN SBovine tuberculosis is one of the major disease affecting humans, bovines and wildlife, by virtue of its economic importance and public health significance. For early diagnosis of the disease, initial attempts were made to genomically characterize the Mycobacterium tuberculosis complex (MTBC) organisms causing disease in bovine or wildlife and to prepare a diagnostic reagent, purified protein derivative (PPD) from one of the more recent strains involved in the disease condition. To ensure adequate quantity and quality of DNA extracted from nasal secretions of animals, cotton or tampon swabs were tested. It was found that tampon swabs could extract more nasal secretion leading to significantly higher DNA yields and quality. The sensitivity of the use of tampon swab in detecting known culture was ten times higher than cotton swabs. The excretory pattern of MTBC and Non tuberculousmycobacteria (NTM) from tuberculin skin test positive animals was erratic and inconsistent. The samples of nasal secretions, milk and lymph node aspirates resulted only in isolation of NTMs over MTBC organisms. The NTM were found to grow abundantly and early as compared to M. bovis isolates. They could probably mask the growth of MTBC leading to preferential isolation of NTMs from bovine excretions especially when the animals are maintained in an environment rich in NTM organisms.ThesisItem Restricted DEVELOPMENT AND CHARACTERIZATION OF DIFFERENT APPROACHES AGAINST DOG ERYTHROCYTE ANTIGEN 1.1 (DEA 1.1)(TANUVAS, CHENNAI, 2022) KALAISELVI G.; TANUVAS; TIRUMURUGAAN K.G.; DHINAKAR RAJ G.; VIJAYARANI, K.; BARANIDHARAN G.R.Dog blood group systems are defined by the expression of various antigens on the surface of red blood cells (RBCs) of which the blood group system DEA I is clinically considered most important due to the strong DEA 1 antigenicity. Hence, accurate typing of canine blood types is essential clinically in repeated transfusions to prevent potentially fatal blood incompatibility reactions. In India, the increased number of hemoprotozoan associated anemia and related transfusions has necessitated indigenous research on different approaches for an attempt to characterize the DEA 1 antigen. Current increasing need for blood transfusion warrants the appropriate identification of the appropriate blood type in dogs (Dog erythrocyte antigen- DEA) which have not been completely characterized as on date. Among these six common reported blood types the DEA l.l is considered as the major antigen type as they are found in more than 60% of the population and to confirm this DEA type there are only three commercially available typing kits (that use the same murine monoclonal antibody) and only one source of dog polyclonal antibody.ThesisItem Restricted BIOHYDROGEN PRODUCTION FROM RUMINAL CONTENTS(2021) Merli Maman; TANUVAS; Meignanalakshmi S; Ghadevaru Sarathchandra; Vijayarani K; Meenakshi SundaramHydrogen gas is a promising alternative for fossil fuel at the current time due to the advantages that is environment friendly, it can be derived from renewable sources and it has high energy efficiency. In the present study, hydrogen gas was producedin a biological way by using rumen fluid of cattle collected from slaughter house by dark fermentation of rumen fluid and Microbial Electrolysis Cell (MEC) process. Integration of both dark fermentation and MEC was also carried out. Dark fermentation of rumen fluid was carried out in 100 ml air tight serum bottles using rumen fluid as the substrate and microorganisms present in the rumen fluid as inoculum and the production of biohydrogen was confirmed by gas chromatography with Thermal conductivity detector (GC-TCD). The dark fermentation parameters such as pH, substrates and micronutrient concentrations were optimized. The pH 4, 5, 6, 7 and 8 were used for dark fermentation of rumen fluid and it was found that pH 6 produced higher biohydrogen when compared to other pH. Substrates such as 1% rice bran, 1% powdered potato peel and 1% powdered tapioca peel were used as added substrates to rumen fluid for dark fermentation and it was found that 1% rice bran produced higher biohydrogen. The micronutrients used were O.25M, 0.5M, 0.75M and IM concentrations of Ferrous sulphate. Magnesium sulpahte and Potassium sulphate. It was found that in all the three mucronutrients, 0,5M concentration produced higher biohydrogen. Rumen fluid collected from slaughter house was used for isolation of biohydrogen producing microorganisms and four different biohydrogen producing bacteria were isolated, characterized and the production of biohydrogen was also confirmed using the individual bacteria in dark fermentation. The rumen fluid after dark fermentation was also given for metagenomic analysis to find out the phylum, genus and species abundance in the rumen fluid. The Microbial electrolysis cell (MEC) was designed and made for the biohydrogen production. The parameters such as pH, salt-bridge, catholyte, electrodes and substrates were optimized for the production of biohydrogen. Among the pH 4, 5, 6, 7 and 8, the highest amount of biohydrogen production was obtained in pH 6. 0.8% and 1% of agarose and agar-agar were used as salt-bridge in the MEC and the highest biohydrogen production was obtained in MEC with 0.8% agarose. The electrodes used were carbons rods and graphite plates where the MEC with graphite plates as the electrode showed higher biohydrogen production. Sodium chloride in different concentrations such as 0.25M, 0.5M, 0.75M and IM were used and it was found that 0.5M Sodium chloride produced highest amount of biohydrogen in MEC. The substrates added to the rumen fluid in the anode chamber were 1% rice bran, 1% powdered potato peel and 1% powdered tapioca peel and the highest amount of biohydrogen production was obtained in the MEC with 1% rice bran.The integration of dark fermentation and MEC was carried out for an enhanced amount of biohydrogen production. The total amount of biohydrogen produced was calculated from the individual amount of biohydrogen produced by dark fermentation and MEC. The quantity of biohydrogenproduced were statistically analysed and the obtained p value is less than 0.005 (p<0.005) which shows that the quantity of biohydrogen produced with different pH and substrates were statistically significant.ThesisItem Restricted DEVELOPMENT OF RECOMBINANT FLAGELLAR PROTEIN BASED IMMUNODIAGNOSTIC ASSAY FOR DETECTION OF Campylobacter jejuni(2021) Suresh, Bobade Sumedha; TANUVAS; Vijayarani, K; Thirumurugan, KG; Thangavelu, A; VairamuthuCampylobacter spp. is a zoonotic pathogen and most reported cause of human gastroenteritis worldwide. Campylobacter jejuni is a commensal organism of the intestinal tract of poultry and other agriculturally important animals. The major transmission routes of Campylobacteriosis in human are consumption of contaminated or undercooked meat, raw milk or improper pasteurized milk. This study was undertaken to investigate the incidence of Campylobacter jejuni in milk, meat and faecal samples. A total of 143 samples comprising mastitis milk (20), raw milk (52), faecal swab of dog (19) suffer from diarrhoea, raw chicken meat (19), meat sample from sheep (5) and goat (3), beef sample (25) were collected aseptically. The samples were studied for cultural, biochemical and molecular examinations.ThesisItem Restricted CLONING AND EXPRESSION OF IMMUNOGENIC ANTIGENS OF Mycobacterium avium subsp. paratuberculosis(2018) Padmavathy, C; TANUVAS; Vijayarani, K; Kumanan, K; Thangavelu, A; Srithar, AParatuberculosis (pTB) is a fatal inflammatory disease caused by Mycobacterium avium subsp. paratuberculosis (MAP) affecting both wild and domestic ruminants.Nospecificand sensitive diagnostic assay is available due to the cross reactivity with the strains of vast mycobacterium family. Development of diagnostics for MAPwill require a cocktail of antigens due to therange of heterogeneous Major histocompatability complex(MHC) molecules and due to differentiated antigen expression patterns of MAP. Recombinant antigens for the Mycobacterium avium subsp. paratuberculosisgenes namely MAP4_0900 encoding 34kDa B cell epitope, MAP specific IS900 - Nir Aencoding T cell epitope, MAP4_ 2856 (ESXO-ESAT) encoding early secretory ESAT 6 like protein,MAP4_1239c encoding virulence factor MmpL were cloned and expressed in E.coli. The gene inserts for the selected genes were amplified byPolymerase chain reaction (PCR) with specific primers designed for site directional cloning. A fusion gene encoding 28 immunogenic peptides of nine genes, Ag85A,Ag85B, Ag85C,P35-35kDa, Serine Protease, AhpC, AhpD, GroE1, Hsp20 was synthesised and obtained as a pUC 57 vector insert.ThesisItem Restricted HEMATOPOIETIC PROGENITOR CELL CHARACTERIZATION IN CANINES(2018) Nandhini, S; Mangala Gowri, A; Meenambigai, TV; Baranidharan, GR; TANUVASBlood cells are responsible for constant maintenance and immune protection of every cell type of the body and this relentless and brutal work requires cells that have the greatest powers of self-renewal and are designated as Hematopoietic progenitor cells (HPCs). Peripheral blood stem cells in circulation have become the most common source of hematopoietic stem cells intended for transplantation after minimal manipulation. In vitro and in vivo animal studies have shown that the enriched CD34+marrow cells of different species can give rise to the multiple blood cell lineages and may provide long-term hematopoiesis, suggesting that CD34 could be regarded as a cell surface marker of primitive hematopoietic stem cells. This has made it possible to enrich HSCs from mice and human beings for molecular characterization and transplantation purposes. Homeobox (Hox), sonic hedgehog (SHH), and Wingless-type MMTV integration site family (Wnt) are known to modulate the self-renewal and expansion of hematopoietic progenitor/stem cells in humans and mice. Unlike cytokines, Hox, SHH, and Wnt are highly conserved among species from flies to humans but studies regarding the self-renewal and expansion of the HSC are extremely limited in dogs. Hence in the present research, a rapid CD34+population enrichment and immuno separation protocol has been standardized in canine blood. The expression of signaling molecules for self renewal and colony forming potential of the isolated CD34+ population of cells from Canine blood showed that they were cells with proliferation and self renewal potential.ThesisItem Restricted EXPRESSING THE INFLUENCE OF EPIDERMAL GROWTH FACTOR AND LEUKEMIA INHIBITORY FACTOR IN THE DEVELOPMENT OF BOVINE PREIMPLANTATION EMBRYOS(2018) Vinayak, Sadekar Gautami; Meenambigai, TV; Mangalagowri, A; Reena, D; TANUVASAn array of growth factors and cytokines are associated in the development of embryos under in vivo and in vitro conditions. These growth factors and cytokines have autocrine and paracrine cell-cell signaling actions that contribute to yielding healthier embryos. Their actions in the embryo include modulation of cell gene expression and metabolic function that in turn influence cell survival and differentiation, ultimately impacting embryo implantation competence and post-implantation development. The effect of Epidermal Growth Factor (EGF) and Leukemia Inhibitory Factor (LIF) in the development of IVM/IVF bovine oocytes /embryos and their gene expression analysis were investigated in this study.
