PB1-F2 Attenuates Virulence of Highly Pathogenic Avian H5N1 Influenza Virus in Chickens

Abstract

Highly pathogenic avian influenza virus (HPAIV) is a permanent threat due to its capacity to cross species barriers and generate severe infections and high mortality in humans. Recent findings have highlighted the potential role of PB1-F2, a small accessory influenza protein, in the pathogenesis process mediated by HPAIV in mammals. In this study, using a recombinant H5N1 HPAIV (wt) and its PB1-F2-deleted mutant (ΔF2), we studied the effects of PB1-F2 in a chicken model. Unexpectedly, when using low inoculation dose we observed that the wt-infected chickens had a higher survival rate than the ΔF2-infected chickens, a feature that contrasts with what is usually observed in mammals. High inoculation dose had similar mortality rate for both viruses, and comparison of the bio-distribution of the two viruses indicated that the expression of PB1-F2 allows a better spreading of the virus within chicken embryos. Transcriptomic profiles of lungs and blood cells were characterized at two days post-infection in chickens inoculated with the wild type (wt) or the ΔF2 mutant viruses. In lungs, the expression of PB1-F2 during the infection induced pathways related to calcium signaling and repressed a large panel of immunological functions. In blood cells, PB1-F2 was associated with a gene signature specific for mitochondrial dysfunction and down-modulated leucocytes activation. Finally we compared the effect of PB1-F2 in lungs of chickens and mice. We identified that gene signature associated to tissue damages is a PB1-F2 feature shared by the two species; by contrast, the early inhibition of immune response mediated by PB1-F2 observed in chickens is not seen in mice. In summary, our data suggest that PB1-F2 expression deeply affect the immune response in chickens in a way that may attenuate pathogenicity at low infection dose, a feature differing from what was previously observed in mammal species.

Influenza A virus PB1-F2 protein prolongs viral shedding in chickens lengthening the transmission window

“…In line with previous results seen investigating functions of PB1-F2 with HPAI viruses (Leymarie et al , 2014), we observed a significant increase in pathogenicity when PB1-F2 was removed from our LPAI H9N2 virus, in an embryonated chicken egg model and in chickens in vivo . We also determined that the presence of PB1-F2 in an H9N2 virus prolonged the infectious shedding period in infected chickens which resulted in an extended transmission window with enhanced transmission.…”

“…Leymaire et al (2014) have previously shown that the presence of a PB1-F2 protein increased the amount of vRNA detected in the intestine of birds inoculated with an HPAI virus at 2 days post inoculation. Using both qRT-PCR for vRNA and infectious virus particle titration, we determined the relative levels of the WT and KO viruses in a panel of respiratory and gastrointestinal tract tissues at both day 2 and 5 post inoculation for the high-dose infection study (Fig.…”

“…Leymarie et al (2014) previously reported that the removal of the PB1-F2 protein from an HPAI H5N1 virus resulted in an increase in chicken mortality at low dose. To determine if the absence of PB1-F2 also increased the pathogenicity of LPAI viruses in chickens, we inoculated groups of chickens via the nares with either a low dose (5×10 3  p.f.u.)…”

“…Two other published studies have undertaken a similar approach to understanding the role of PB1-F2 in the context of avian species infection with influenza viruses (Leymarie et al , 2014; Schmolke et al , 2011). However, both focused solely on the effect that a full-length PB1-F2 protein has on the pathogenic nature on H5N1 viruses which were already highly pathogenic.…”

“…Work to delineate the function of avian influenza virus PB1-F2 in avian hosts has only been limited to highly pathogenic H5N1 influenza strains and these studies have concentrated on reading out pathogenic outcomes (Hulse-Post et al , 2007; Leymarie et al , 2014; Schmolke et al , 2011). No publications have examined the role of PB1-F2 in a LPAI background, which make up the vast majority of avian influenza infections globally.…”