Highly pathogenic or low pathogenic avian influenza virus subtype H7N1 infection in chicken lungs: small differences in general acute responses

Abstract

Avian influenza virus can be divided into two groups, highly pathogenic avian influenza virus (HPAI) and low pathogenic avian influenza virus (LPAI) based on their difference in virulence. To investigate if the difference in clinical outcome between LPAI and HPAI in chickens is due to immunological host responses in the lung within the first 24 hours post infection (hpi), chickens were infected with LPAI or HPAI of subtype H7N1. Virus was found in the caudal and cranial part of the lung. With LPAI, virus was localised around the intrapulmonary bronchus and secondary bronchi. In sharp contrast, HPAI was detected throughout the whole lung. However, based on viral RNA levels, no quantitative difference was observed between LPAI and HPAI infected birds. In infected areas of the lungs, an influx of CD8α+ cells as well as KUL01+ macrophages and dendritic cells (DC) occurred as fast as 8 hpi in both infected groups. No major difference between LPAI and HPAI infected birds in the induction of cytokines and interferons at mRNA level in lung tissue was found.In conclusion, the differences in lethality for chickens infected with LPAI or HPAI could be ascribed to difference in location of the virus. However similar amounts of viral RNA, similar cytokine mRNA levels, and similar influxes of CD8α+ and KUL01+ macrophages and DC were found between HPAI and LPAI in the lungs. A cytokine storm at mRNA level as described for mammals was not observed in the lungs of HPAI infected birds within 24 hpi.

Figure 1

Scatter plot of the amount of viral RNA of individual birds in the tracheal swabs at different time points post inoculation (hpi) with LPAI (open symbols) or HPAI (closed symbols). The horizontal line represents the mean of six individual birds. Viral RNA expression was determined using RT-PCR and data were expressed as EID₅₀/g.

Figure 2

Scatter plot of the amount of viral RNA of individual birds in the lung at different time points post inoculation (hpi) with LPAI (open symbols) or HPAI (closed symbols). The horizontal line represents the mean of 6 individual birds. Viral RNA expression was determined using RT-PCR and data were expressed as EID₅₀/g.

Figure 3

Detection of AIV (nucleoprotein positive cells) in the lung by immunohistochemistry at 24 hpi (A and C) and 16 hpi (B and D). LPAI infected cells are localised around the intrapulmonary bronchus and in adjacent parabronchi (A), but not in the periphery (B). HPAI infected cells are detected around the intrapulmonary bronchus (C) and in peripheral parabronchi (D).

Figure 4

Detection of CD8α+ (A-C) and KUL01+ (D-E) cells in lung 16-24 h after AIV infection. In LPAI infected chickens influxes of CD8α+ cells are mainly detected in and adjacent to the intrapulmonary bronchus (A) and rarely in peripheral parabronchi (B). In HPAI infected chickens influxes of CD8α+ are also detected in peripheral parabronchi (C). Influxes of KUL01+ cells were localised in and adjacent to the intrapulmonary bronchus after infection with LPAI (D), whereas KUL01+ cells are detected in peripheral parabronchi after infection with HPAI (E). These cellular influxes co-localised with virus infected cells.

Figure 5

Pro-inflammatory cytokine mRNA expression in the cranial and caudal part of the lung after inoculation with LPAI (white bar) or HPAI (black bar) at different time points post inoculation (hpi). IL-1β, IL-6 and IL-8 mRNA expression was shown as fold induction compared to uninfected birds ± SD. * p < 0.05 between infected and control chickens; line p < 0.05 between the LPAI and HPAI group of a specific time point pi.

Figure 6

Interferon mRNA expression in the cranial and caudal part of the lung after inoculation with LPAI (white bar) or HPAI (black bar) at different time points post inoculation (hpi). IFN-α, IFN-β and IFN-γ mRNA expression was shown as fold induction compared to uninfected birds ± SD. * p < 0.05 between infected and control chickens; line p < 0.05 between the LPAI and HPAI group of a specific time point pi.