1 B-5064 [1-6] RESEARCH ARTICLE Indian Journal of Animal Research Molecular and Immunopathological Investigations of Marek’s Disease in HVT Vaccinated Chicken Flocks Shailja Pandey 1 , Deepika Lather 1 , Aman Kumar 2 , Vikas Nehra 1 , Babu Lal Jangir 1 , Gulshan Narang 1 10.18805/IJAR.B-5064 ABSTRACT Background: The present investigation was carried out on poultry carcasses brought to the Department of Veterinary Pathology, LUVAS, Hisar from Herpesvirus of turkeys (HVT) vaccinated chicken flocks and suspected for viral neoplastic conditions on the basis of necropsy. Conventional and molecular techniques for the confirmation of the Marek’s disease (MD) were studied. For differential diagnosis from other oncogenic viral infections immunohistochemistry as an important tool was investigated. Methods: The conventional diagnosis of MD) was done on basis of clinical signs, gross lesions and histopathology of proliferate lesions; as lymphomatous lesions consist of pleomorphic lymphocytes. For confirmatory diagnosis molecular techniques were employed to detect MD virus specific Meq gene by Taqman probe real time PCR and nested PCR. For differential diagnosis of mixed oncogenic viral infections, immunohistochemistry (IHC) using B and T cell differentiation markers (CD79 alpha and CD3) was employed. Result: Specific Meq gene by Taqman probe real time PCR was positive in 92.5% suspected cases; while nested PCR technique gave positivity in 55% cases. The immunohistochemical reactivity for CD3 gave positive reactivity in the cytoplasm and cell membrane and negative reaction for CD79 alpha in MD positive cases, while in cases with mixed infections of MDV and other viral neoplastic conditions, cells showed positive reactivity to both CD3 and CD79 alpha in neoplastic proliferates. Key words: CD79 alpha and CD3, Immunohistochemistry, Marek’s disease, Nested PCR, Real time PCR. INTRODUCTION Marek’s disease virus serotype 1 (MDV1) is an oncogenic DNA virus which belongs to the family Herpesviridae subfamily Alphaherpesvirinae and genus Mardivirus causing the Marek’s disease (MD) of poultry. Major broader clinical signs in MD are immunosuppression, polyneuritis and lymphoproliferative nodules formation in the visceral and cutaneous tissues (Witter and Schat, 2003). The traditional diagnosis of MD is based on clinical signs and gross pathology. Histopathological examination of proliferate lesions is also very important in diagnosing MD; as lymphomatous lesions consist of pleomorphic lymphocytes. Even with the intensive vaccination practice, outbreaks of MDV associated lymphocytic tumors and paralysis was reported from India (Kamaldeep, 2007) and other countries resulting in great economic losses. Additionally, the increasing virulence and the immunosuppressive effect of MDV makes it a threat to broiler industry as it has been reported to increase their susceptibility to Escherichia coli and infectious bronchitis (IB) infection (Islam et al., 2002). Prompt and differential disease diagnosis therefore plays a key role to minimise the losses by enabling early adaptation of proper control and preventive measures. The principal methods to confirm the avian oncogenic diseases are isolation of the virus, demonstration of viral DNA or antigens in tissues and detection of antibody (Schat and Nair, 2008). PCR appears to be a method of choice for the diagnosis of avian oncogenic viruses and IHC can be used in differential diagnosis of these neoplastic conditions (Gopal et al., 2012). Therefore; for optimizing production, it is mandatory to use the correct pathological, microbiological and other diagnostic techniques, which may be helpful to check the disease occurrence and minimize the losses in production. In the present study, we report the diagnostic investigation of MD in HVT vaccinated organized poultry farms of Haryana using immunopathohistochemical and molecular techniques. 1 Department of Veterinary Pathology, College of Veterinary Sciences, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar-125 004, Haryana, India. 2 Department of Animal Biotechnology, College of Veterinary Sciences, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar-125 004, Haryana, India. Corresponding Author: Deepika Lather, Department of Veterinary Pathology, College of Veterinary Sciences, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar-125 004, Haryana, India. Email: drdeepika@luvas.edu.in How to cite this article: Pandey, S., Lather, D., Kumar, A., Nehra, V., Jangir, B.L. and Narang, G. (2023). Molecular and Immunopathological Investigations of Marek’s Disease in HVT Vaccinated Chicken Flocks. Indian Journal of Animal Research. doi:10.18805/IJAR.B- 5064. Submitted: 05-12-2022 Accepted: 22-04-2023 Online: 09-06-2023