A set of RT-PCR assays for detection of all known avian paramyxoviruses and application in surveillance of avian paramyxoviruses in China.

BACKGROUND: Avian paramyxoviruses (APMVs), also termed avian avulaviruses, are of a vast diversity and great significance in poultry. Detection of all known APMVs is challenging, and distribution of APMVs have not been well investigated. METHODS: A set of reverse transcription polymerase chain reaction (RT-PCR) assays for detection of all known APMVs were established using degenerate primers targeting the viral polymerase L gene. The assays were preliminarily evaluated using in-vitro transcribed double-stranded RNA controls and 24 known viruses, and then they were employed to detect 4,346 avian samples collected from 11 provinces. RESULTS: The assays could detect 20-200 copies of the double-stranded RNA controls, and detected correctly the 24 known viruses. Of the 4,346 avian samples detected using the assays, 72 samples were found positive. Of the 72 positives, 70 were confirmed through sequencing, indicating the assays were specific for APMVs. The 4,346 samples were also detected using a reported RT-PCR assay, and the results showed this RT-PCR assay was less sensitive than the assays reported here. Of the 70 confirmed positives, 40 were class I Newcastle disease virus (NDV or APMV-1) and 27 were class II NDV from poultry including chickens, ducks, geese, and pigeons, and three were APMV-2 from parrots. The surveillance identified APMV-2 in parrots for the first time, and revealed that prevalence of NDVs in live poultry markets was higher than that in poultry farms. The surveillance also suggested that class I NDVs in chickens could be as prevalent as in ducks, and class II NDVs in ducks could be more prevalent than in chickens, and class II NDVs could be more prevalent than class I NDVs in ducks. Altogether, we developed a set of specific and sensitive RT-PCR assays for detection of all known APMVs, and conducted a large-scale surveillance using the assays which shed novel insights into APMV epidemiology.

Phylogenetic Analyses of Fowl Adenoviruses (FAdV) Isolated in China and Pathogenicity of a FAdV-8 Isolate.

Fowl adenoviruses (FAdVs) have a worldwide distribution and are associated with a variety of diseases, causing considerable economic losses to the poultry industry. We characterized 10 FAdVs isolated from China in 2015-2016 and assessed the pathogenicity of a FAdV-8 strain in specific-pathogen-free (SPF) chickens. Phylogenetic analysis of a hexon gene revealed that only 1 of the 10 isolates belonged to FAdV-8, whereas others belonged to FAdV-4, indicating that Chinese FAdVs were mainly FAdV-4 in recent years. The pathogenicity experiment of the FAdV-8 strain CH/SD/2015/09 showed that no clinical signs were observed in infected chickens. Necropsy displayed mild necrotic foci and petechial hemorrhage of livers collected at 5 days postinfection (dpi). Histopathologic examination identified the presence of intranuclear inclusion bodies in hepatocytes. No virus was detected in oral and cloacal swabs at 5 dpi, and only viral DNA could be measured in kidneys collected at the same time. The results revealed that CH/SD/2015/09 had no obvious pathogenicity in 5-wk-old SPF chickens, which could provide a better understanding about the pathogenicity of the FAdV-8 serotype.

Different Origins of Newcastle Disease Virus Hemagglutinin-Neuraminidase Protein Modulate the Replication Efficiency and Pathogenicity of the Virus.

To investigate the exact effects of different origins of Newcastle disease virus (NDV) hemagglutinin-neuraminidase (HN) protein to the biological characteristics of the virus, we systematically studied the correlation between the HN protein and NDV virulence by exchanging the HN of velogenic or lentogenic NDV strains with the HN from other strains of different virulence. The results revealed that the rSG10 or rLaSota derivatives bearing the HN gene of other viruses exhibited decreased or increased hemadsorption (HAd), neuraminidase and fusion promotion activities. In vitro and in vivo tests further showed that changes in replication level, tissue tropism and virulence of the chimeric viruses were also consistent with these biological activities. These findings demonstrated that the balance among three biological activities caused variation in replication and pathogenicity of the virus, which was closely related to the origin of the HN protein. Our study highlights the importance of the HN glycoprotein in modulating the virulence of NDV and contributes to a more complete understanding of the virulence of NDV.