Abstract
Since the first report from Cherry Valley ducks on a commercial duck farm in China (2008), duck astrovirus type 1(DAstV-1) -associated duck viral hepatitis (DVH) have been detected in several commercial duck flocks. In the literature, no outbreak of DAstV-1 have been report in China since 2012. Here, the isolation, cultivation and characterization of DAstV-1 isolate are described. One DAstV-1 strain, designated as DAstV-SDZZ, was isolated from a diseased duckling. The isolated astrovirus grew well in the LMH cell line. To determine the entire genomic of the DAstV-SDZZ isolate, next-generation sequencing (NGS) technique was conducted on Illumina HiSeq platform. Complete genome sequence analysis revealed that DAstV-SDZZ isolate was 91.6%-98.7% homology with others DAstV-1 deposited in Genbank. Similar clinical symptoms were successful reproduced by experimental infection study using the DAstV-SDZZ isolate. DAstV-SDZZ is the first DAstV-1 strain whose experimental infection study has been conducted. The present works are likely to provide new insights into the pathogenicity and evolution of DAstV-1 in ducks.
Keywords: duck astrovirus type 1; next-generation sequencing; complete gene sequence; phylogenetic analysis; pathogenicity
Introduction
Astroviruses are non-enveloped, positive-sense and single-stranded RNA viruses with a diameter of approximately 28–30 nm (Baxendale and Mebatsion, 2004; Fauquet et al., 2005). Their single-stranded RNA genome ranges in size from 6.8 to 7.9 kb and consisting of three open reading frames (ORFs) ; ORF 1a, ORF 1b and ORF 2 (Fu et al., 2008; De Benedictis et al., 2011). ORF1a codes for non-structural (NS) polyprotein, which include a serine protease, several transmembrane (TM) helical motifs and a nuclear localization signal (NLS) (Jiang et al., 1993; Lee and Kurtz, 1994). ORF 1b encodes the RNA-dependent RNA polymerase (Willcocks et al., 1994), while ORF2 codes for the viral capsid structural protein that is required for virion formation (Arias and Dubois, 2017).
In poultry, astroviruses have been reported to infect several avian species, including chickens (Kang et al., 2018), turkeys (Pantin-Jackwood et al., 2006), gooses (Q. Zhang et al., 2018; Yang et al., 2018) and ducks (Chen et al., 2012; Wei et al., 2020). Astroviruses disease in most species are causative agents of enteric diseases. In ducks, unlike other avian species, astroviruses have been associated with a fatal hepatitis, historically known to be caused by two types of duck hepatitis virus (DHV-2 and DHV-3) which were considered picornaviruses (Asplin, 1965; Haider and Calnek, 1979). More recently, DHV-2 and DHV-3 has been identified as astrovirus based on a 391 nt ORF 1b region (Todd et al., 2009). Besides the two types of formal classification duck astrovirus (DAstV), there are a number of unassigned duck astrovirus waiting to be classified, such as duck DHV-3-like astroviruses (Liu, Wang, Shi et al., 2014), duck astrovirus CPH (DAstV CPH) (Liao et al., 2015), duck astrovirus YP-like and astroviruses detected from wild birds (Chu et al., 2012; Liu, Wang and Zhang, 2014).
In China, duck viral hepatitis is mainly caused by duck hepatitis A virus (DHAV) (Wen et al., 2018). No outbreaks of DAstV-1 have been published reports in China since 2012 (Chen et al., 2012). Therefore, little is known regarding their molecular biology and viral genetic diversity. In addition, due to the lack of effective methods for cultivating DAstV-1, the pathogenicity of the DAstV-1 in ducklings remains unclear. The present paper describes the molecular characterization of this virus and phylogenetic analysis to other astroviruses. The reproduction experiment was conducted to investigate the pathogenicity of the virus.