Molecular characterization of velogenic viscerotropic Ranikhet (Newcastle) disease virus from different outbreaks in desi chickens

Abstract

Aim: Diagnosis of velogenic viscerotropic Ranikhet disease from six different flocks of desi chicken in and around Mumbai by gross and histopathological examination, isolation of virus and molecular methods.

Materials and methods: A total of 25 carcasses (varying between 2 and 6 carcasses from each flock) of six different flocks of adult desi chicken were subjected to necropsy examination for diagnosis of the disease during the span of a year (2014-2015). After thorough gross examination, the tissue samples were collected and processed for virus isolation and histopathological examination. The 20% tissue homogenate was inoculated into 9-day-old specific pathogen free (SPF) embryonated eggs. Mean death time (MDT) of embryos after inoculation and intracerebral pathogenicity index (ICPI) were used to judge velogenic nature of the virus. Newcastle disease virus (NDV) was isolated from six cases and confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) targeting the partial fusion protein gene of the viral genome.

Results: A total of 25 carcasses (varying between 2 and 6 carcasses from each flock) of six different flocks of desi chicken were presented for postmortem examination to Department of Veterinary Pathology, Bombay Veterinary College, Parel, Mumbai during 2014-2015. The gross and histopathological examination revealed lesions suggestive of viscerotropic velogenic form of the Newcastle disease (ND). The 20% tissue homogenate was inoculated into 9-day-old embryonated eggs from SPF chicken. NDV was isolated from six cases and confirmed by RT-PCR targeting the partial fusion protein gene. MDT of all the isolates was <60 h which indicated velogenic nature of the virus. ICPI of the isolates ranged between the 1.63 and 1.78. In four out of six outbreaks concurrent moderate to heavy infection of Ascardii galli in one flock and Railetina spp. in three flocks was also noted. In this study, viscerotropic velogenic form of ND was confirmed in all six outbreaks by gross and microscopic examination, virus isolation and RT-PCR.

Conclusions: In this study, viscerotropic velogenic form of ND was confirmed in all six outbreaks by gross and microscopic examination, virus isolation and RT-PCR. Nowadays, vaccine strains Lasota, B1 and F strains are used widely in India to control the infection of NDV. However, virulent NDV strains are still isolated frequently in the birds under backyard and also in commercial venture which demonstrates that NDV remains an on-going threat to commercial as well as backyard poultry flocks.

Keywords: Newcastle disease virus; mean death time; reverse transcriptase polymerase chain reaction; specific pathogen free; velogenic.

Figure-1

Gross picture showing hemorrhages at the tip of proventricular glands (arrows with up and down heads).

Figure-2

Hemorrhagic ulcers in intestine seen through serosa (arrow with down head).

Figure-3

Hemorrhages in cecal tonsils (arrow with up head).

Figure-4

Section of intestine showing extensive hemorrhages (arrow with down head), multifocal necrosis (arrow with up head), loss of epithelial cells lining villi and infiltration of inflammatory cells (arrow with horizontal head). (H and E, 100×).

Figure-5

Section of trachea with necrosis of tracheal epithelial cell along with mild infiltration of inflammatory cells (arrow with up head), congestion of vessels and edema in submucosa. (H and E, 100×).

Figure-6

Reverse transcriptase polymerase chain reaction amplification of the fusion protein gene of RD virus (1148 bp product) from three cases.