Abstract
Although wild ducks are considered to be the major reservoirs for most influenza A virus subtypes, they are typically resistant to the effects of the infection. In contrast, certain influenza viruses may be highly pathogenic in other avian hosts such as chickens and turkeys, causing severe illness and death. Following in vitro infection of chicken and duck embryo fibroblasts (CEF and DEF) with low pathogenic avian influenza (LPAI) viruses, duck cells die more rapidly and produce fewer infectious virions than chicken cells. In the current study, the morphology of viruses produced from CEF and DEF cells infected with low pathogenic avian H2N3 was examined. Transmission electron microscopy showed that viruses budding from duck cells were elongated, while chicken cells produced mostly spherical virions; similar differences were observed in viral supernatants. Sequencing of the influenza genome of chicken- and duck-derived H2N3 LPAI revealed no differences, implicating host cell determinants as responsible for differences in virus morphology. Both DEF and CEF cells produced filamentous virions of equine H3N8 (where virus morphology is determined by the matrix gene). DEF cells produced filamentous or short filament virions of equine H3N8 and avian H2N3, respectively, even after actin disruption with cytochalasin D. These findings suggest that cellular factors other than actin are responsible for the formation of filamentous virions in DEF cells. The formation of elongated virions in duck cells may account for the reduced number of infectious virions produced and could have implications for virus transmission or maintenance in the reservoir host.
Highlights
Virus replication following infection of chicken and duck cells was measured at 2–48 h post infection by titration of infectious virus using a focus forming assay in Madin Darby canine kidney cells (MDCK) cells
Virus production was comparable between species up to 8 h pi, but after 24 h and 48 h pi there was a significant difference in virus production between species, with chicken cells producing 4–5 fold more virus than duck cells (Fig. 1a)
Replication of H2N3 virus in chicken and duck cells was confirmed by measuring matrix protein 1 (M1) gene ribonucleic acid (RNA) and protein expression following cell infection with the virus for 8 and 24 h, using quantitative real-time polymerase chain reaction (PCR) and western blotting, respectively
Summary
Influenza A viruses show variable morphology, with shapes ranging from spherical or elliptical and about 100 nm in diameterto elongated or filamentous with a length reaching to more than several micrometres; occasionally, they are pleomorphic (Calder et al, 2010). Influenza A viruses show variable morphology, with shapes ranging from spherical or elliptical and about 100 nm in diameter. There is a matrix protein 1 (M1) layer. All these proteins play an important role in virus morphogenesis (Bouvier and Palese, 2008; Palese and Shaw, 2007). Diversity of virus morphology is thought to be a genetic trait; in particular the seventh viral RNA segment (M), which encodes the matrix proteins, plays a dominant role in determining virus shape (Elleman and Barclay, 2004; Roberts et al, 1998). The importance of specific M protein residues as determinants of virus morphology appears to differ between influenza viruses of different species (Elton et al, 2013). The surface glycoproteins (HA and NA) have been implicated in modulation of virus shape (Jin et al, 1997; Zhang et al, 2000)
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