Detection of Marek’s disease virus meq gene in Feather follicle by Loop-mediated Isothermal Amplification

Abstract

Marek's disease, caused by lymphotropic alphaherpesvirus, is one of the most economically significant diseases of poultry. Despite the ubiquitous use of vaccination to control losses, Marek's disease still affects poultry farming worldwide with an estimated annual loss up to US $2 billion. There is a need for a simple and rapid diagnostic method for early detection of Marek's disease virus from clinical samples under field conditions. A loop-mediated isothermal amplification assay was developed using a set of six primers in order to detect meq gene segment of oncogenic Marek's disease virus (serotype-1) in feather tips of chickens. Bst polymerase was used for the isothermal amplification of viral DNA at 65 o C for 60 min in a water bath. The fluorescence signal was identified in Marek's disease virus-positive samples after the addition of SYBR Green I under ultraviolet illumination. The loop-mediated isothermal amplification technique was found to be more sensitive as compared to Marek's disease virus-specific PCR for the detection of Marek's disease caused by oncogenic Marek's disease virus. The results demonstrate a significant advantage of loopmediated isothermal amplification in diagnosis of Marek's disease compared to conventional PCR as it can be carried out in most situations where rapid diagnosis is required, like in field conditions.

Marek's disease herpesvirus serotype 1 in broiler breeder and layer chickens in Malaysia

“…MD has been causing significant economic losses to the poultry industry as a result of the mortality and morbidity that it causes in poultry, particularly, chicken. These economic losses have been estimated to be about 2 billion USD annually [9]. The economic loss due to MD is further increased as the current vaccination-based control and prevention strategy is being undermined due to the continued enhancement of the pathogen virulence.…”

“…MD is a major economic risk for poultry farms as it occurs in almost all commercial chicken farms and causes significant economic loss with an estimated annual loss up to the US $2 billion worldwide [8,9]. Mortality rarely exceeds 10-15% and can occur over a few weeks or many months [10].…”

Identification and Characterization of Marek’s Disease Virus Serotype 1 Using Molecular Approaches

“…Analisis hasil uji LAMP dapat dilakukan dengan spektrofotometer, penggunaan pewarna fluorescent atau diintegrasikan dengan mesin loopamp realtime turbidimeter yang mampu untuk mengukur intensitas fluorescent yang dikorelasikan dengan konsentrasi amplikon DNA yang teramplifikasi. Beberapa penelitian dengan metode LAMP telah dilakukan dalam rangka mendapatkan uji identifikasi virus MDV-1 yang bersifat murah dan mudah diaplikasikan pada kondisi lapangan (Woźniakowski et al 2011b;Angamuthu et al 2012;Lawhale et al 2014;Woźniakowski & Samorek-Salamonowicz 2014). Selanjutnya, Woźniakowski et al (2013) mengembangkan uji LAMP untuk membedakan tiga serotipe virus Marek, yaitu MDV-1, GaHV3 dan HVT.…”

Clinical, Pathological and Molecular Study of Marek’s Disease in the Gallus gllus Domesticus and BrahmaChickens of Basra Province, Iraq

“…There are several diagnostic methods for MD, clinical signs, post mortem changes, histopathological changes and the use of polymerase chain reaction (PCR) [6]. MD is a major economic risk for poultry farms as it occurs in almost all commercial chicken farms and causes significant economic loss with an estimated annual loss up to the US $2 billion worldwide [7]. Due to the unpredictability of outbreaks and the possibility of vaccination failure as a consequence of the evolution of more virulent strains of MDV, MD remains a major concern for the poultry industry [8].…”