Pathak, Debesh ChandraBEHERA, DEBASISH2022-07-012022-07-012019-07https://krishikosh.egranth.ac.in/handle/1/5810185068The present research work was carried out with an aim to study the pathology necrotic enteritis with isolation and molecular detection of C. perfringens and experimental production of the NE in chicken to compare between the C. perfringens type A and C in terms of hematological, biochemical and pathomorphological alterations. Total 320 numbers of samples based on different clinical signs/pathological conditions were collected from 15 districts of Assam. Isolation and identification of C. perfringens was done by cultural and morphological characteristics and confirmation was done by detection of cpa gene of C. perfringens by PCR. In this study 20 (15.03%) intestinal content and 9 (4.81%) cloaca swabs were found to be positive for cpa gene of C. perfringens. Isolation of bacteria from the samples collected during monsoon was found to be highest in comparison to other seasons. Study showed around 80% of the total isolates of C. perfringens was from the birds of 4-6 weeks of age. The C. perfringens isolated from the enteritis samples were found to be highest. Total 29 samples were found to be positive for cpa gene (324 bp) encoding for alpha gene and cpb gene (180 bp) encoding for beta gene was detected in 11 isolates. The additional virulence toxin genes of C. perfringens like TpeL and NetB were also detected. The gross lesions of NE in field condition revealed haemorrhagic, eroded, detached dead mucosal tissues in intestine. Formations of diphtheritic membrane, distention of intestine were also observed in intestine. Liver, kidneys and lungs showed congestion, haemorrhage and focal areas of necrosis. Spleen and Bursa of Fabricious in some birds was found to be moderately enlarged. The gross lesion of brain was found to be limited to mild congestion of meningeal blood vessels. Histopathology of NE in chickens revealed congestion of blood vessels in the lamina propria and submucosa with vacuolation of epithelial cells of intestinal villi along with necrosis making the villi broader and shorter. Different developmental stages of coccidia were also seen in the mucosal epithelial cells. In other organs such as liver, kidneys, heart, lungs, spleen, bursa of Fabricius and brain showed variable nature of histopathological lesions like congestion, haemorrhage and focal areas of coagulative necrosis. Experimental production of NE was done in chicken by infecting C. perfringens isolate Type A and type C with and without coccidia in separate groups. The clinical signs shown by the experimentally infected birds were diarrhoea, dehydration, depression, reluctance to move, loss of appetite, ruffled feathers, drooping of wings and head and huddling. The clinical pathology of experimental birds showed, significant decrease in TEC level, hemoglobin (g/dl) level as well as in PCV (%) and significant increase in TLC in the birds of infected group in comparison to the control. Serum ALT and AST both showed a significant increase (P<0.01) and total protein showed a significantly decreased (P<0.05) level. The gross lesion revealed congestion and haemorrhage and focal areas necrosis of mucosa of intestine. Enlargement, congestion, haemorrhage with focal areas of necrosis of the liver were common gross findings in all the experimentally infected groups. This might be due to damage to RBC in entero-hepatic circulation by α toxin of Clostridium perfringens. Kidneys, heart, lungs, spleen and Bursa of Fabricius revealed moderate degree of congestion and haemorrhage. Variable degrees of vascular changes in terms of gross lesions were observed in all most all the infected groups. The histopathological lesions revealed developmental stages of coccidia (schizonts & merozoites) and infiltration of large no of mononuclear cells and few polymorhonuclear cells in intestine. The intestinal villi have undergone necrosis and necrosed cells were sloughed off from the mucosa. Liver revealed marked fatty change in hepatocytes, congestion in the sinusoids. Kidneys showed focal areas of inter tubular congestion. Heart and lungs revealed focal areas of mononuclear cell infiltration as well as congestion and haemorrhage. Spleen and Bursa of Fabricius showed depletion of follicles and brain showed neuronal degeneration and necrosis with neuronophagia. Based upon the clinical signs, gross and histopathology, the present study revealed the groups infected by both coccidia and clostridial isolate showed distinctly more pronounced qualitatively and quantitatively in terms of clinical signs and pathological lesions. It has been also observed in this study that C. perfringens type A was found to be more virulent in terms of pathogenesis and pathomorphology in comparison to C. perfringens type C. TEM evaluation of experimentally infected birds showed disruption of intercellular junctional complexes, formation of gaps between enterocytes and delimitation of boundaries of individual enterocytes. Disintegration of nuclear material, dilatation of endoplasmic reticulum, disruption of cristae of mitochondria, increase intracytoplasmic vacuolizations and membrane bound vesicles were also prominent ultrastructural alterations in this study. Data were subjected to statistical analysis and analyzed by SAS System ('Local', X64_7PRO) using one way analysis of variance (ANOVA). Means were presented as mean ± standard error (SE) and were compared by the Duncan test at the 0.05 level of probability.EnglishThesis