Ana Chumbe,^a Ray Izquierdo-Lara,^a Luis Tataje Lavanda,^a Aling Figueroa,^a Karen Segovia,^b* Rosa Gonzalez,^b Giovana Cribillero,^b Angela Montalvan,a Manolo Fernández Díaz,^a Eliana Icocheab. Farvet S.A.C. Chincha Alta, Ica, Perúa. Laboratory of Avian Pathology, Facultad de Medicina Veterinaria, Universidad Nacional Mayor de San Marcos (UNMSM), Lima, Perúb. * Present address: Karen Segovia, Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA.

ABSTRACT

Here, we report the first complete sequence and biological characterization of a Newcastle disease virus (NDV) isolated from a peacock in South America (NDV/peacock/Peru/2011). This isolate, classified as genotype XII in class II, highlights the need for increased surveillance of noncommercial avian species.Newcastle disease virus (NDV) is able to infect250 species of birds worldwide and is also responsible for an acute and very contagious respiratory disease. NDV (genus Avulavirus, family Paramixoviridae) possesses a negative-sense single-stranded RNA genome. Wild birds have been implicated in the spread of the disease to domestic birds (1). Based on sequence analysis of the fusion protein (F) gene, NDV strains are classified in class I or II. Strains of class I are avirulent in chickens, whereas strains in class II can be lentogenic, mesogenic, or velogenic. Class II is subdivided into at least 18 genotypes from I to XVIII (2). Several outbreaks have been observed in Peru in recent years, but information regarding which genotypes are circulating is poor. In 2011, 9 out of 10 peacocks (Pavo cristatus) died in a zoo farm in Huachipa, Lima, Peru, with nervous and respiratory symptoms.

The virus isolation, from oral and cloacal swabs, was carried out in 9-day-old embryonated specific-pathogen-free (SPF) chicken eggs. Allantoic fluid (AF) was collected and assessed by hemagglutination and a hemagglutination inhibition test using NDV hyperimmune serum. Amplified AF was clarified, aliquoted, stored at 80°C, and quantified by plaque assay. The 50% lethal dose (LD₅₀) was calculated in 4-week-old SPF chickens by 10-fold serial dilutions.

The intracerebral pathogenicity index (ICPI) was calculated according to standard protocols (3). All protocols that included animals were approved by the Institutional Animal Care and Use Committee of the Universidad Nacional Mayor de San Marcos (UNMSM), Lima, Peru. The complete sequence of NDV/peacock/Peru/2011 was amplified by 8 pairs of overlapping oligonucleotide primers using the OneStep reverse transcription-PCR (RT-PCR) kit (Qiagen). Purified amplicons were cloned into the pGEM-T Easy (Promega) and sequenced with T7/SP6 primers.

The assembly and alignment were made on SnapGene and Clustal W, respectively. Phylogenetic analysis and sequences comparison were done in MEGA 6.0.

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