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ThesisItem Open Access PATHOMORPHOLOGICAL STUDIES ON ENTERITIS IN SLAUGHTERED SHEEP(SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2022-07) MONIKA, K; SUDHA RANI CHOWDARY, CH(MAJOR); RAMA DEVI, V; SREEDEVI, CIn the present study, a total of 395 sheep from slaughter houses in and around Vijayawada and field mortalities were screened for enteritis. Of these, enteritis was recorded in 110 sheep with an occurrence of 27.84%. Based on gross and histopathological examination, enteritis was classified into catarrhal (20.9%), haemorrhagic (7.3%,), necrotic (9.1%) and parasitic enteritis (62.7%). The etiological agents identified were parasites viz. immature amphistomes, Schistosoma spp., Moniezia expansa, Moniezia benedeni, Stilesia globipunctata, Trichuris ovis, Oesophagostomum spp. and Eimeria spp., bacteria viz. E. coli, Salmonella spp., and Clostridium perfringens and a virus, PPRV. Catarrhal enteritis was of acute and chronic types characterized by presence of catarrhal contents in the lumen of intestines, goblet cell hyperplasia and infiltration of neutrophils and lymphocytes in the lamina propria in acute type, and mononuclear cells in chronic type. Haemorrhagic enteritis was characterized by presence of blood mixed contents, severe mucosal congestion and haemorrhage, coagulative necrosis, villus atrophy and fusion, infiltration of lamina propria with a large number of mononuclear cells and neutrophils, and presence of Gram positive bacteria. Necrotic enteritis was characterized by severe coagulative necrosis of the intestinal mucosa, fusion and atrophy of villi, lacteal dilation, severely hyperplastic to necrotic and cystic crypts, infiltration of the lamina propria with a large number of mononuclear cells and presence of Gram positive bacteria. Majority of the cases of necrotic enteritis were associated with PPR virus infection which showed severe congestion to petechial haemorrhages along the mucosal folds of ileo-caecal junction and rectum giving a zebra striped appearance, syncytial cells in the intestinal mucosa and lymphocytolysis in the Peyer’s patches. In the present study, parasitic enteritis included immature amphistomiasis, schistosomosis, monieziosis, stilesiosis, trichuriosis, oesophagostomosis and coccidiosis. Grossly, in cases of immature amphistomiasis, monieziosis, stilesiosis, and trichuriosis, the parasites were observed in the lumen of intestines, while, schistosomosis, oesophagostomosis and coccidiosis were diagnosed on microscopic examination of the intestines. Nodular lesions in the intestines were observed grossly in schistosomosis, stilesiosis and oesophagostomosis which on histopathology were found to be egg granulomas, hyperplastic villi and pseudotubercular granulomas respectively. In cases of coccidiosis, small greyish white nodules were observed in the intestines which microscopically were found to be hyperplastic crypts and villi harbouring different stages of Eimeria spp. In the present study, 73 samples of intestines were subjected to bacteriological studies. E. coli was the major bacteria isolated from cases of catarrhal enteritis, immature amphistomiasis, monieziosis, stilesiosis, trichuriosis and coccidiosis. Clostridium perfringens was isolated from cases of haemorrhagic enteritis, necrotic enteritis, immature amphistomiasis, monieziosis and coccidiosis, while, Salmonella spp. was isolated only from cases of necrotic enteritis. In the present study, intestines and mesenteric lymph node samples of PPR suspected animals were subjected to one step RT-PCR using a set of primers targeting the N gene of PPRV and yielded a product of 350 bp fragment of N gene in seven samples confirming the presence of PPR virus. In conclusion, the results of the present study revealed occurrence of different types of enteritis, their pathomorphologies and associated etiological agents.ThesisItem Open Access PATHOMORPHOLOGICAL AND MOLECULAR STUDIES OF CANINE TRANSMISSIBLE VENEREAL TUMOR(SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2022-06) YASHASWI, YALAVARTHI; SAMATHA, V(MAJOR); RAMA DEVI, V; RAVI KUMAR, PIn the present study, a total of 72 dogs were suspected for canine TVT based on history and clinical signs. Samples from all the suspected cases were collected from the Veterinary Clinical Complex, NTR College of Veterinary Science, Gannavaram, Veterinary polyclinics, private pet clinics and animal birth control centres present in the coastal districts of Andhra Pradesh. Canine TVT was diagnosed in 68 dogs based on cytology, histopathology, histochemistry, IHC, transmission electron microscopy and molecular studies with an overall occurrence of 94.4%. Canine TVT in dogs was recorded in six coastal districts of Andhra Pradesh and the highest occurrence of TVT was found in Guntur district (100%), followed by Krishna (95.4%), East Godavari (91.6%), West Godavari (90.9%), Visakhapatnam (90%) and Prakasam district (87.5%). The occurrence of canine TVT in the present study was highest in the winter (45.6%), followed by the rainy (29.4%) and summer seasons (25%). High occurrence of TVT was seen in dogs aged between 2-4 years (72.1%), dogs aged 4-6 years (16.2%), 6-8 years (8.8%) and >8 years (2.9%). The occurrence of canine TVT was notably higher in female dogs (83.8%) when compared to that of males (16.2%). In the present study, canine TVT was recorded mostly in non-descript dogs when compared to other breeds. The occurrence of tumor location in 57 female dogs was recorded and was highest at the vestibulo-vulval junction (47.4%), vulval lips (19.3%), vulval mucous membrane (17.5%), posterior vagina (15.8%) and six cases revealed tumors on extragenital sites like skin at perineal region in addition to genital tumors. In 11 male dogs, the occurrence of venereal tumor growth was highest at the base of the penis (36.4%), prepuce (36.4%) and on the tip of penis (27.2%). One case showed tumor at extra genital site on the skin in inguinal area in addition to the genital tumor. Most of the TVT suspected dogs showed clinical signs like bloody discharges from genital tracts in both sexes, excessive licking of genitalia, deformation of external genitalia and perineal region due to large tumors. Grossly, TVTs appeared as pink fleshy or dark firm tumors. Tumors on genitalia of both sexes at initial stages appeared as small hyperaemic papules and the progressing tumors appeared nodular, papillary or multilobulated as well as caulifower-like or pedunculated tumors. The predominant cytomorphological pattern of TVT in the present study is the plasmacytic type, followed by the lymphocytic and mixed types. Cytological examination of 68 cases out of 72 suspected dogs revealed canine TVT cells. Canine TVTs were classified into three cytomorphological sub types: plasmacytic (53%), lymphocytic (28%) and mixed (19%), based on the type of tumor cells. Plasmacytic type tumor cells were large ovoid cells with abundant cytoplasm and several clear cytoplasmic vacuoles at cell margins. The cartwheel-shaped nucleus was eccentrically placed and showed one or more prominent nucleoli in the majority of cells. Lymphocytic tumors cells were round cells with scanty cytoplasm. A high nuclear to cytoplasm ratio was evident in the majority of cells. A few cells revealed cytoplasmic and nuclear vacuolation. The nucleus was round and hyperchromatic with coarse chromatin and prominent nucleoli. Mixed type of tumor cells revealed both lymphocytic and plasmacytic tumor cells. Binucleated cells, mitotic figures, neutrophils, erythrocytes and macrophages were observed in many smears. The cytological smears collected from extragenital TVTs in the cutaneous region revealed polymorphonuclear cells and keratinized squamous cells along with TVT cells. In the present study, 68 tissue samples from genital tumors revealed characteristic histopathological features of TVT. The tissue sections revealed a higher number of neoplastic plasma cells and lymphocytes. Histologically, TVT sections were categorised into progressive (35), steady (12) and regressive stages (21). Progressing tumors contained densely packed tumor cells with scanty connective tissue stroma and a significantly higher number of mast cells, mitotic figures and microvessels at the invasion of the tumor. Mild proliferation of fibroblastic cells along with loosely packed round tumor cells were observed in the steady stage of the tumor. The tumors in the regression phase had decreased round tumor cells, decreased mitosis, increased infiltration of TILs and apoptotic cells. Increased proliferation of fibroblasts and increased collagen was evident. Tissue sections (7) from the extra genital tumor growth on the skin had numerous round neoplastic cells in the dermis. The proliferative cellular activity in TVT cells was demonstrated by AgNOR staining in 68 TVT sections. The mean values of AgNOR count were high in progressing tumors (> 7 AgNOR dots) and low in steady (3-7 AgNOR dots) and regression tumors (< 3 AgNOR dots). Immunohistochemical studies of TVT for PCNA and vimentin were carried out. Proliferating cell nuclear antigen (PCNA) showed strong to moderate expression in progressive tumors (61.8%) and in regressing tumors (38.2%) respectively. Vimentin is a specific marker of mesenchymal derivation and immunoreactivity to vimentin was intense in the cytoplasm of more than 90% of the tumor cells. Four TVT positive tissues were subjected to transmission electron microscopy and two revealed plasmacytic, one lymphocytic and one mixed type. Loosely distributed tumor cells and closely packed tumor cells with small protoplasmic microvilli and extensive interdigitation with adjacent cells were observed. Neoplastic lymphocytes and plasma cells were observed and the lymphocytes were round cells with central oval to irregularly round nucleus whereas, plasmacytes were large oval shaped cells with vacuolated cytoplasm and prominent RER with numerous ribosomal clusters. Initial phases of tumor growth revealed elongated cells with elongated nuclei and presence of neovascularization. The presence of more nucleoli and big perichromatin granules were observed, indicating mitotic figures in progressing tumor sections. Regressing tumors revealed collagen between the tumor cells, fibroblast like cells, tissue infiltrating leucocytes and apoptotic cells. Genomic amplification of 22 TVT tissue samples for demonstration of rearrangement of the c-myc gene segment using specific MycS-2 and LINE AS-1 primers revealed an expected product of 550 bp. Based on the cytology, histopathology, histochemistry, immunohistochemistry, electron microscopy and molecular studies, tumors from 68 cases were canine transmissible venereal tumors consisting of lymphocytic, plasmacytic and mixed type of cells at different stages of tumor development.ThesisItem Open Access PATHOMORPHOLOGICAL STUDIES ON SPONTANEOUS KIDNEY LESIONS IN SLAUGHTERED PIGS(SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2022-06) PADMINI, VAKA; HEMANTH, I(MAJOR); RAMA DEVI, V; SRINIVASA RAO, TThe present study was undertaken to know the occurrence of various spontaneous kidney lesions in slaughtered pigs, to describe the pathomorphological features of kidney lesions and to identify the probable etiological agents. A total of 320 slaughtered pigs of either sex were screened for various pathological conditions of kidney, if any, at different locations in Krishna District, Andhra Pradesh. Gross and microscopic kidney lesions of pathological significance were observed in 130 animals (40.6%) that were grouped under circulatory disturbances (13.85%), degenerative changes (6.92%), inflammatory conditions (60%), parasitic (11.54%) and miscellaneous (7.69%) conditions. Circulatory disturbances comprised of congestion (10.77%) and haemorrhages (3.08%). Renal congestion was characterized by dark red coloured kidneys with engorged blood capillaries in the cortex and medulla. Renal haemorrhages revealed diffuse petechial and large ecchymotic haemorrhages on the surface with numerous extravasated RBCs in the interstitium and within the tubules. Nephrosis refers to the degenerative changes in the renal parenchyma, was characterized by enlarged and pale kidneys with cloudy swelling and vacuolar degeneration of tubular lining epithelium along with the formation of protein and cellular casts in the tubules. Nephritis was categorized into glomerulonephritis (26.93%), interstitial nephritis (32.31%) and tubulo-interstitial nephritis (0.77%). Glomerulonephritis was comprised of acute (3.08%), sub-acute (21.54%) and chronic (2.31%) types. Acute lesions were characterized by pale and edematous kidneys with petechiae on the cortex along with glomerular lesions of congestion, swelling, hypercellularity and reduced Bowman’s space with protein accumulation. Sub-acute type revealed enlarged and pale kidneys with epithelial cresents and synechiae formation, sclerotic glomeruli, peri-glomerular mononuclear cells infiltration and perivasculitis. Chronic type revealed smaller sized kidneys with finely granular surface and lesions of diffuse global glomerular shrinkage, atrophy and necrosis with severe fibroplasia and mononuclear cells infiltration. Interstitial nephritis was comprised of sub-acute (29.23%) and chronic (3.07%) types. Sub-acute condition was characterized by variable sized whitish foci on the renal surface with nodular and diffuse aggregation of lymphoplasmacytic cells around the tubules, glomeruli and blood vessels. In chronic cases, kidneys were hard with diffuse pale areas and revealed diffuse and extensive proliferation of connective tissue all over the interstitium along with severe mononuclear cells infiltration. Tubulo-interstitial nephritis (0.77%) revealed kidneys with greyish white foci surrounded by hyperemic rim and lesions of multi-focal tubular necrosis and liquefaction with infiltrating polymorphs and mononuclear cells, and diffuse interstitial mononuclear cells infiltration. Stephanurosis (kidney worm) affections were characterized by numerous parasitic nodules in the perirenal fat, renal pelvis and around the ureter harbouring the adult and larval forms of the parasite. Microscopically, thick fibrous nodules containing cross section of parasites and severe infiltration of plasma cells, lymphocytes, macrophages and giant cells in the nodule wall were noticed. In the present study, 7.69% of renal cysts was recorded under miscellaneous conditions. Renal cysts were characterised by clear cystic cavities in the renal parenchyma with clear watery fluid with thick limiting membrane. Microscopically, empty or fluid filled cavities lined by single layer of flattened epithelial lining surrounded by proliferative connective tissue proliferation were noticed. Etiological studies revealed isolation of pathogenic bacterial species in 20% samples that comprised of Staphylococcus spp., Streptococcus spp., Escherichia coli and Klebsiella spp. in association with renal congestion, nephrosis, nephritis and kidney worm affections. The present study concluded that a significant occurrence of spontaneous renal lesions exist in slaughtered pigs and the most common pathological condition encountered being nephritis with consistent lesion of mononuclear cell infiltration. Further, this study suggests that a detailed investigations by involving large sample size and employing different molecular detection methods for wider range of etiological agents including zoonotic importance if any, so as to implement proper preventive measures in pig farming and to minimise the economic losses and to curtail zoonotic spill over, in any.ThesisItem Open Access PATHOLOGICAL AND MOLECULAR DIAGNOSIS OF INFECTIOUS BURSAL DISEASE IN COMMERCIAL CHICKEN(SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2022-03) AKHILA, N; SATHEESH, K (MAJOR); SAMATHA, V; SRINIVASA RAO, TThe present study was undertaken to know the occurrence of IBD in commercial chicken in around Vijayawada and to carry out pathological and molecular diagnosis of IBD. In the present study, a total of 922 IBD suspected birds of 4 to 6 weeks age were procured from different places in and around Vijayawada, from private diagnostic laboratories besides postmortems conducted in NTR College of Veterinary Science, Gannavaram. At necropsy, IBD was diagnosed in 252 birds out of 922 screened with an overall occurrence of 27.33% based on pathological lesions and molecular detection of IBDV by RT-PCR. The disease was more prevalent in 4 to 6 weeks aged birds with majority of cases occurring during the 5th week (40.11%). The occurrence was highest in April to September months (31 to 40%). As per the history, the affected birds showed depression, watery diarrhoea (white to yellowish), soiled vents, anorexia, ruffled feathers, reluctance to move, closed eyes and death. Necropsy conducted in 252 birds revealed predominant gross lesions in the bursa of Fabricius, spleen, thymus, caecal tonsils. Petechiae and ecchymoses on the pectoral and thigh muscles, haemorrhages at the proventriculus-gizzard junction. Bursa was swollen and oedematous with creamy or yellowish discoloration, gelatinous exudates in the serosa, congestion and haemorrhages on the serosal and mucosal surfaces and inner surface of longitudinal laminae in almost all the cases. Occasionally, the bursa became atrophied and whitish or creamy, which sometimes contained cheesy mass in the lumen. The spleen was enlarged and mottled. Kidneys were markedly swollen, congested and liver was enlarged, friable and yellowish in few cases. Histopathology revealed the prominent findings in bursa of Fabricius, characterized by congestion and hemorrhages along with lymphoid depletion of various degrees in the follicles resulting in atrophy of follicles and formation of cystic spaces within the follicles. Presence of eosinophilic necrotic material in the bursal follicles along with degenerating heterophils, atrophy of plicae, widening of interfollicular spaces with proliferation of fibrous connective tissue and sloughing of bursal epithelium was observed in few cases. Microscopic examination of spleen depicted congestion of splenic vessels and sinusoids, depletion of lymphocytes of varying degrees in white pulp and germinal centers. Thymus revealed decrease in thymocyte density in the cortex and congestion in most of the cases. However, in some cases disruption of normal architecture of cortex and medulla was also evident. Lymphoid depletion in lymphoid nodules, mucosal necrosis along with infiltration of mononuclear cells in the lamina propria were noticed in caecal tonsils in majority of the cases. Kidneys revealed interstitial haemorrhages and congestion in peritubular capillaries, severe infiltration of inflammatory cells, degeneration and individualization of tubular epithelial cells with detachment from the basement membrane and glomerular shrinkage. Liver revealed haemorrhage and severe necrosis of hepatocytes, congestion in the central vein, fatty change in hepatic parenchyma, diffuse coagulative necrosis of hepatocytes, along with severe infiltration of mononuclear cells in the portal areas of the liver. In the present study, pooled bursal samples were collected from 33 birds and utilized for molecular diagnosis of by RT-PCR using specific primers targeting VP2 gene of IBDV. Electrophoretic analysis revealed an amplified product of 480 bp fragment of VP2 gene in 9 samples thus confirming the presence of IBDV. The nucleotide sequencing and phylogenetic analysis of present IBDV sample (OM2024620) revealed it’s close relation with the strains of Kerala (MK 861022.1), Gujarat (AB605259.1), Karnataka (EU788042.1) and Haryana (AY739668.1). In the present study, the pathological and molecular diagnosis of IBD was carried out and the occurrence of IBD in commercial chicken in and around Vijayawada was recorded. It was evident that IBD continues to be a major threat causing economic losses in chicken in the study area
