Impact of chicken-origin cells on adaptation of a low pathogenic influenza virus

Abstract

Understanding the growth dynamics of influenza viruses is an essential step in virus replication and cell-adaptation. The aim of this study was to elucidate the growth kinetic of a low pathogenic avian influenza H9N2 subtype in chicken embryo fibroblast (CEF) and chicken tracheal epithelial (CTE) cells during consecutive passages. An egg-adapted H9N2 virus was seeded into both cell culture systems. The amount of infectious virus released into the cell culture supernatants at interval times post-infection were titered and plaque assayed. The results as well as cell viability results indicate that the infectivity of the influenza virus was different among these primary cells. The egg-adapted H9N2 virus featured higher infectivity in CTE than in CEF cells. After serial passages and plaque purifications of the virus, a CTE cell-adapted strain was generated which carried amino acid substitutions within the HA stem region. The strain showed faster replication kinetics in cell culture resulting in an increase in virus titer. Overall, the present study provides the impact of cell type, multiplicity of infection, cellular protease roles in virus infectivity and finally molecular characterization during H9N2 virus adaptation procedure.

Comparative Transcriptomic Analysis of Immune-Related Gene Expression in Duck Embryo Fibroblasts Following Duck Tembusu Virus Infection

“…IRF7, is activated by TLR-dependent signaling pathways and it regulates type I IFN responses [44]. NF-κB is a vital transcription factor that regulates genes responsible for a variety of immune response [42].…”

Isolation and characterization of a distinct duck-origin goose parvovirus causing an outbreak of duckling short beak and dwarfism syndrome in China

“…If the embryo died, the allantoic fluid and internal organs were tested for waterfowl parvovirus antigens by LA and IFA. Duck embryo fibroblasts (DEF) cells were prepared from 11-day-old Cherry Valley duck embryos as described previously [19]. DEF monolayer cell cultures were inoculated with allantoic fluid, and cultures were incubated at 37°C in 5 % CO 2 for 7 days, followed by three further passages in DEF.…”

Variability of tropism and replicative capacity of two naturally occurring influenza A H9N2 viruses in cell cultures from different tissues

“…It has been shown that difference among cell tropism may be due to different amounts of endogenous trypsin-like enzyme present in the cells that subsequently activate the cleavability of the HA. It was demonstrated that recent H9N2 viruses possess di-and tribasic HA cleavage sites RXXR (the case of our variants) or RSK/RR motifs (Aamir et al, 2007;Xu et al, 2007) which are activated by type II transmembrane serine proteases (TTSPs) (Baron et al, 2013), HAT (human airway trypsin-like protease), TMPRSS2 (transmembrane protease, serine S1 member 2), expressed in the human airway epithelium, the gastrointestinal, and urogenital tracts (Bottcher et al, 2013) and VAP and chTMPRSS13 in the chicken tracheal epithelium cells (Bertram et al, 2010a(Bertram et al, , 2010bShahsavandi et al, 2013). Likewise, another cellular protease, matriptase, which is expressed in the epithelial cells of most tissues but with higher expression levels in the kidney (Baron et al, 2013), was able to support proteolytic activation of H9N2 viruses showing cleavage site motifs R-S-S-R or R-S-R-R, but not those containing a single arginine or motif A-S-R-R.…”