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Birsa Agricultural University, Ranchi

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Subject: Veterinary Pathology × End date: 2019 × Type: Thesis × Thesis Type: Ph.D ×
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    ThesisItemOpen Access
    Studies on Pathology of Fenvalerate intoxication in Goats with Special Reference to Immunopathology
    (Birsa Agricultural University, Ranchi, 1992) Kaushal Kumar Singh; K.P. Yadava
    Chemical pesticides are playing the pivotal role in augmenting the crop production throughout the world. Injudicious use of pesticides in the field and on animal body has culminated in toxicity and altered our ecosystem. Thus in a bid to evolve non-toxic pesticides, synthetic pyrethroids have been discovered with greater safety of mammalian species due to its quick degradation in the ecosystem. However, a safety evaluation of any pesticide should include a clear understanding of its mode of toxic action on different organs so that a preventive and curative measure may be brought into books in order to minimise the risk posed by the pesticides in our ecosystem or on animal body. Keeping this point in view, two experiments were designed to test the short and long term toxic effect of fenvalerate in goats. In experiment-I seven young Black Bengal male goats were procured and divided on at random basis in two groups viz. control and treated group. The treated group of four animals were drenched with a single dose of fenvalerate at the rate of 400 mg/kg body weight and clinical, biochemical and morpholo gical studies were carried out. The clinical signs of the acute toxicity consisted of grinding of teeth, opisthotonus, excessive salivation incoordination, prostration, coms and death. There was a significant increase in serum AST and ALT after three hours but serum LDH showed increase in its value after six hours of drenching. On the other hand, no significant alteration in the level of bun could be observed at any stage of the acute toxicity.
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    ThesisItemOpen Access
    Study of Cypermethrin intoxication in Goats with Special Reference to immunopathology
    (Birsa Agricultural University, Ranchi, 1995) Madhrendu Kumar Gupta; G. J. Jha
    Chemical pesticides have become an integral part of modern agricultural practices. Extensive use of pesticides to augment farm production has increased the risk of health hazards to man and animals and hence it is necessary to know every facets of these toxicants so that suitable preventive and curative measures could be advocated to the users in particular and grazing animals in general. The slow biodegradation of certain pesticides poses greater risk to biological life and hence all the missing gaps in our knowledge must be bridged without being complacent. We have used goat model to study the short and long term effect of chemical pesticide. For the purpose, two sets of experiments were designed viz. acute and chronic toxicity experiment. To study the acute toxicity, ten young, growing Black Bengal male goats were randomly divided into two groups such as control and intoxicated. Intoxicated group consisted of six goats whereas control group had only four animals. Intoxicated animals were drenched with a single dose of cypermethrin at the rate of 500 mg/kg body weight and clinical signs of the toxicity including biochemical alterations, if any, were studied. The clinical signs of the acute cypermethrin intoxication consisted of nervous manifestations such as staggering gait, posterior weakness followed by lateral recumbaney, opisthotonus, tonic spasm and shivering. In addition, there was grinding of teeth, excessive salivation, dyspnoea followed by coma and death. There was a significant progressive increase in serum ALT, AST, blood urea and serum LDH level. Post mortem examination of the dead animals revealed congestion and haemorrhage in most of the visceral organs such as liver, heart, lunge, kidney and meninges of the brain. Mild hydropericardium was also noticed. On the other hand, micro scopic examination of liver revealed wide spread vacuolar degeneration of hepatocytes and mild centrilobular necrosis. In addition, there was congestion of sinusoida, portal vein and hepatic arteries. In kidney there was acellular and eosino philic albuminous precipitates in the tubular lumen and vacuolar degeneration alongwith congestion was seen in the glomeruli and in the intertubular capillaries. Satellitosis, neuronophagia and mild perivascular cuffing were seen in the cerebrum. Extensive degenerative changes were recorded in the seminiferous tubules of the testes. To find out the long term effect of cypermethrin on goats a chronic toxicity experiment was designed. Twelve young, growing Black Bengal male goats were divided into two groups. The intoxicated group had seven goats and they were daily drenched with cypermethrin at the rate of 30 mg/kg body weight for a period of 90 days. Chronic toxicity symptoms appeared only after 20 days of intoxication. There was development of post erior weakness, roughness of hair coat, loss of body weight, dullness, depression and lacrymation. It is significant to note that the symptoms appeared after drenching of the pesticide and lasted for 2 hours. Biochemical estimations revealed significant increase in the level of serum ALT, AST, AP and blood urea. On the contrary, decrease was noticed in the level of serum AchE act ivity, total protein, albumin and globulin. However, no signi ficant variation was observed in the level of serum calcium and inorganic phosphorus. The effect of chronic cypermethrin toxicity on humoral immunity was assessed by monitoring th
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    ThesisItemOpen Access
    Pathology and pathogenesis of Clostridium perfringens type- D epsilon Toxin in Goats
    (Birsa Agricultural University, 1986) Parhi, Mobin Kumar; Chauhan, H.V.S.
    Studies on "The pathology and pathogenesis of Clostridium perfringens type D epsilon (6) toxin in goats" were conducted because of prevalence of the disease in goats in South Bihar and because many cases must have gone undiag nosed. Also the roles of epsilon toxin one of the main toxic products of this organism were poorly understood. The studies were conducted on natural cases of enterotoxaemia due to cl. perfringens type D in goats as well as in goats inoculated with crude (epsilon) toxin, and purified epsilon toxin in high, medium and low doses. The studies on goats revealed characteristic symptoms like, convulsions, opisthotonus, salivation, bulging of eyballs and glycosuria etc. The main, diagnostic gross pathological alterations were "pulpy kidneys" due to nephrosis, thickening of interlobular septae of anterior lobes of the lungs due to oedema, endocardialhaemorrhages, haemorrhagic spots in brain and increased amount of fluid in small intestine. Encephalograms of intoxicated goats revealed status epilepticus in all the experimental groups. Cardiograms revealed arrhythmia, atrial fibrillation, ventricular premature beats, increased amplitude and flutter in goats given high dose of epsilon toxin but not in those given medium and low doses.
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    ThesisItemOpen Access
    USE OF ACRIDINE ORANGE STAINING FOR SIMULTANEOUS ASSESSMENT OF PROLIFERATIVE AND APOPTOTIC ACTIVITY IN N-NITROSO-N-METHYLUREA INDUCED TUMOURS IN RATS
    (Birsa Agricultural University, Kanke, Ranchi, Jharkhand, 2015) Kumar, Kaushal; Singh, K.K.
    Biological activity of a tumour is determined by a balance between proliferative and apoptotic activity of its neoplastic cells. Histopathological typing and grading has remained a gold standard to assess proliferative aggressiveness of a tuomour which relies mostly on anaplastic and apoptotic changes shown by neoplastic cells.But many times these changes do not reveal true proliferative behavior of a tumour. These problems motivated the augmentation of histopathological methods to improve the accuracy of assessment of proliferative and apoptotic activity of tumours. Proliferative behavior of a tumour is determined by mean growth fraction and mean cycle time of their neoplastic cells. Growth fraction of tumour in a tissue section is assessed by measurement of proliferation associated antigens like PCNA, Ki-67 etc. But no technique is Abstract……… Page 2 available to see the speed of proliferation of neoplastic cells particularly in terms of mean cycle time instantaneously in tissue section of tumours. This highlights the importance of identification and quantification of cells in different phases of cell cycle to throw more insight on proliferative speed and fraction of neoplastic cells. Different protocols of flow cytometry for identification and quantification of neoplastic cells in different phases of cell cycle in isolated cell system are available using acridine orange (AO) staining. These methods can not reveal complete picture of location, migration, differentiation and relation of dividing cells with other cells in their true histological architecture. So in this study an attempt was made to establish a protocol to identify, localize and quantify such cells in microscopic sections of different NMU induced tumours in Wistar rats. Moreover, AgNORs are the only parameter used to assess proliferative fraction as well as proliferative speed of a tumour on a tissue section indirectly. It has the advantage that their numbers are increased only in actively and fastly dividing cells. The major problem in acceptance of this method as an indirect marker of proliferative speed in terms of mean cycle time is deposition of innumerable fine dots in nuclei in background of large AgNOR dots which has been greatly variable from cell to cell depending upon their metabolic and proliferative status, most possibly in different phases of cycle. Hence, critical analysis of variation in pattern of deposition of such small dots in nuclei has scope to evolve as a system for identifying cells in different phases of cycle. This motivated to evolve one more new system of identifying and localizing dividing cells in different phases of cell cycle on the basis of variation in pattern of deposition of such Non-AgNOR dots in nuclei in histological sections after AgNOR staining. The procedure for optimally demonstrating highest intensity of fluorescence of DNA and RNA along with cellular characteristics for necrotic, apoptotic and neoplastic cells passing through different phases of cell cycle as described and accepted by the method of flow cytometry was accepted and being described as follows. For acridine orange staining of formalin fixed paraffinzed tissue sections of tumours were treated with 0.025% AO in 1 % acetic acid for one minute and then were washed in M/15 Phosphate Buffer (pH 6.0) for few second and sections were mounted in a drop of phosphate buffer and used for examination under fluorescent microscope to identify and count different types of cells. Abstract……… Page 3 The characteristic feature accepted for identification of cells in different phases of cell cycle was based on variation in the colour of fluorescence given by nucleus and cytoplasm of the cell. Dull green fluorescing nuclei and cytoplasm was characterized as cell in G1 phase whereas dull green nuclei with red cytoplasm was characterized as late G1 phase of cell cycle. The cells with bright green to white yellow nuclei with orange cytoplasm were identified as S phase. S/G2 phase of the cell cycle was characterized by bright green to yellow-white nuclei with red fluorescing cytoplasm. The cells with bright green nuclei with mild or no red cytoplasm were identified as M phase of the cell cycle. Apoptotic cells were characterized by condensed orange to red nuclei with red cytoplasm whereas cells with homogeneously fragmented red fluorescing nuclei were identified as necrotic cell. These characteristic feature was accepted for identification of cells by AO staining was highly comparable with those well accepted for flow cytometry taken as standard for identification of such cells in this study. However identification of cells in histological section by AO staining have advantage over flow cytometry that it gave complete picture of apoptotic activity of tumours in terms of their number, location, migration, differentiation and architectural arrangements of dividing cells. One more new system to identify and localize proliferating cells in histological sections was also evolved using AgNOR staining after a critical perusal of variation in pattern of formation and distribution of non- AgNORs along with their nature of persistent reproducibility and ability to be dissolved in sodium thiosulphate solution in AgNOR stained sections of different tumours. The characteristic features incorporated in this system were based on formation and distribution pattern of AgNOR dots and non-AgNOR dots in the nucleus. G1 phase was characterized as cells having large AgNOR dots with few small or medium sized non- AgNOR dots. The Cells with innumerable very fine and small non-AgNOR dots with or without large AgNOR dots inside nucleus was in S phase. G2 phase was characterized as cells having small to medium sized innumerable non-AgNOR dots with brown backgrounds whereas cells with black dense irregular hairy nucleus was considered to be in M phase. The cells in S, G2, and M phases of cell cycle were collectively considered to be in proliferative phase. The accuracy of both these systems to identify, localize and quantify neoplastic cells in different phases of cell cycle was further verified by seeing a correlation between counts of such Abstract……… Page 4 cells in histologic sections of different types of NMU induced tumours after AgNOR as well as AO staining. Keeping these facts in mind, both these systems were further validated by seeing a correlation between counts of cells in different phases of cycle after AgNOR staining and percentage count of quiescent, proliferative and apoptotic cells after AO staining with pathomorphological aggressiveness of different types of NMU induced tumours in wistar rats assessed in terms of tumour volume, multinodularity, total cell count and their impact on survival period. Out of 60 female virgin Wistar rats of about 35 days of age were intoxicated with NMU at the dose rate of 50mg/kg body weight intraperitoneally at 50th, 57th, 64th and 71st days of their age; In order to obtain complete pathomorphological behavior of NMU induced tumour, all the experimental animals were allowed to survive till they died either due to growth of tumour or due to NMU toxicity which prolonged for one year. A total of 44 tumours could be induced in 32 tumour bearing rats. Tumour incidence was found to be 53.33%. The average number of tumours per animal (tumour frequency) was found to be 1.33. The average survival period was 276 days. The mean value of nodularity was found to be 3.25 among the NMU induced tumours. The average volume of the tumour was found to be 26.18 cm3. These induced tumours were further classified into five histologycally distinct types which were mucinous adenocarcinoma (10), tubular adenocarcinoma (7), stromal cell sarcoma (8), mammary ductular adenocarcinoma (12) and gastric squamous cell carcinoma (7) which revealed almost their characteristic histopathological features. Count of cells in different phases of cell cycle after AO and AgNOR staining in histologic sections of almost all these different types of NMU induced tumours were found to be highly and positively correlated and thus comparable with one another. It supported the validity of both these systems to identify cells in different phases of cell cycle. Sensitivity of both these systems were further verified indirectly by correlating the count of cells in different phase of cell cycle with that of proliferative aggressiveness of all these different types of NMU induced tumours as assessed by their volume, total number of cells, multinodularity, grading and survival period which were mostly found to be positively and significantly correlated favouring the ability of this system to identify cells in different phases of cycle. Abstract……… Page 5 A critical perusal of variation in number of cells in different phases of cycle and thereby relative timing taken by neoplastic cells in different phases failed to show a general trend of variation in different histologic types of tumours. Rather, number of cells in different phases of cycle was greatly variable in individual tumour to tumour. It warrants the count of such cells in individual tumour, the critical analysis of which may throw more insight on their proliferative behavior. Higher count of cells in S-phase, G2 phase as well as in M-phase indicated presence of more actively proliferative population and actively growing tumour. Such findings can also suggest that such tumours will give more response with chemotherapy since these agents act mostly on replicating cells A simultaneous count of cells undergoing apoptosis along with count of cells in different phases of cycle will help not only in assessing proliferative behavior of tumour rather it may be of great help in assessing sensitivity or resistance of tumour to a particular drug because different drug acts through different mechanism, which involved inhibition of cells in different phases of cycle or by inducing apoptosis. Mutational changes in these loci may be responsible for a refractory of a tumour to different therapeutic agents. These protocols have a potential to evolve as a test to help in selection of most appropriate chemotherapeutic agent against a particular tumour. Thus it could be concluded that both these systems are helpful in identifying and counting of cells in different phases of cell cycle along with number of cells in apoptosis and can help in throwing more insight on proliferative fraction as well as proliferative speed of a tumour and relative timings in different phases of their cell cycle. Such assessment has got immense importance in prognostic, diagnostic and therapeutic decisions on tumour. However, both these systems of counting cells in different phases of cycle or undergoing apoptosis have still many scopes for improvements as well as exploration to make it more accurate reliable and usable in other areas of tumour and cell biology.