Protection patterns in duck and chicken after homo- or hetero-subtypic reinfections with H5 and H7 low pathogenicity avian influenza viruses: A comparative study

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Abstract

Avian influenza viruses are circulating continuously in ducks, inducing a mostly asymptomatic infection, while chickens are accidental hosts highly susceptible to respiratory disease. This discrepancy might be due to a different host response to the virus between these two bird species and in particular to a different susceptibility to reinfection. In an attempt to address this question, we analyzed, in ducks and in chickens, the viral load in infected tissues and the humoral immune response after experimental primary and secondary challenge infections with either homologous or heterologous low pathogenicity avian influenza viruses (LPAIV). Following homologous reinfection, ducks were only partially protected against viral shedding in the lower intestine in conjunction with a moderate antibody response, whereas chickens were totally protected against viral shedding in the upper respiratory airways and developed a stronger antibody response. On the contrary, heterologous reinfection was not followed by a reduced viral excretion in the upper airways of chickens, while ducks were still partially protected from intestinal excretion of the virus, with no correlation to the antibody response. Our comparative study provides a comprehensive demonstration of the variation of viral tropism and control of the host humoral response to LPAIV between two different bird species with different degrees of susceptibility to avian influenza. © 2014 Chaise et al.

Figures

  • Figure 1. Design of experimental infections of ducks and chickens. 1animals were primo-infected by intranasal and oropharyngeal routes with 4.106 PFU of H5N3 virus, 2animals were primo or reinfected with 4.106 PFU of H5N3 (in red) or H7N2 viruses (in blue), 3OS = Oropharyngeal Swabs, 4CS = Cloacal Swabs, 5Sero = Serology. doi:10.1371/journal.pone.0105189.g001
  • Figure 2. Viral excretion in upper airways (left) or in lower digestive tract (middle) and antibody response (right) to LPAIV H5N3 in ducks (top) or in chickens (bottom). Viral load in oropharyngeal swabs (A, D) and cloacal swabs (B, E) was expressed as viral RNA copies per sample and compared between control (naı̈ves) and challenged birds two days (for chickens, corresponding to the peak of infection in lung) or three days (for ducks, corresponding to the peak of infection in cloacum) after inoculation. All the swabs were eluted in 1.5 ml PBS and were analysed using strictly the same protocol for RNA extraction and RT-PCR. Antibody titration (C, F) was expressed as inverse dilution of serum used for measurement by ELISA (vertical scores) or haemagglutination inhibition method (HI, draught-board). doi:10.1371/journal.pone.0105189.g002
  • Figure 3. Viral excretion in upper airways (left) or in lower digestive tract (right) of LPAIV H5N3 in ducks (top) or in chickens (bottom). Birds were either primo-infected at six weeks of age (H5 groups) or re-infected at six weeks of age with the same virus as that used three weeks before (H5H5 groups). Viral titrations were measured at 2 days (d2, for chickens), 3 days (d3, for ducks) or 8 days p.i. (d8, for both bird species) and were expressed as viral RNA copies per sample and compared between primo-infected (H5 groups, black bars) and re-infected birds (H5H5 groups, white bars). All the swabs were eluted in 1.5 ml PBS and were analysed using strictly the same protocol for RNA extraction and RT-PCR. Significant differences between groups are indicated with asterisks (*, P,0.05; **: P,0.01; ***: P,0.001). doi:10.1371/journal.pone.0105189.g003
  • Figure 4. Antibody response to the H5N3 virus in ducks (top) or in chickens (bottom). Antibody titers were measured by ELISA (left) or HI method (right) from serum collected at the indicated time in days (d) after primary infection (H5 groups, black bars) or secondary infection (H5H5 groups, white bars) and were expressed as the reciprocal of the dilution of serum used for measurement as indicated in the Materials and Methods section. Significant differences between groups are indicated with asterisks (*, P,0.05; **: P,0.01; ***: P,0.001). doi:10.1371/journal.pone.0105189.g004
  • Figure 5. Viral excretion in lower digestive tract of ducks (A) or in upper airways of chickens (B) of LPAIV H7N2. Birds were either primo-infected at six weeks of age (H7 groups) or re-infected at six weeks of age with H7N2 virus, three weeks after H5N3 first inoculation (H5H7 groups). Viral titrations were measured 2 days (d2, for chickens) in oropharyngeal swabs, 3 days (d3, for ducks) in cloacal swabs, or 8 days (d8, for both birds species in their respective swabs) and were expressed as viral RNA copies per sample and compared between primo-infected (H7 groups, black bars) and re-infected birds (H5H7 groups, white bars). All the swabs were eluted in 1.5 ml PBS and were analysed using strictly the same protocol for RNA extraction and RT-PCR. Significant differences between groups are indicated with asterisks (*, P,0.05; **: P,0.01; ***: P,0.001). doi:10.1371/journal.pone.0105189.g005
  • Figure 6. Antibody response to LPAIV H7N2 in ducks (top) or in chickens (bottom). Antibody titers were measured by ELISA (left) or HI method (right) from serum collected at the indicated time in days (d) after primary infection (H7 groups, black bars) or heterologous secondary infection (H5H7 groups, white bars) and were expressed as the reciprocal of the dilution used for measurement as indicated in the Materials and Methods section. Significant differences between groups are indicated with asterisks (*, P,0.05; **: P,0.01; ***: P,0.001). doi:10.1371/journal.pone.0105189.g006

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Chaise, C., Lalmanach, A. C., Marty, H., Soubies, S. M., Croville, G., Loupias, J., … Guérin, J. L. (2014). Protection patterns in duck and chicken after homo- or hetero-subtypic reinfections with H5 and H7 low pathogenicity avian influenza viruses: A comparative study. PLoS ONE, 9(8). https://doi.org/10.1371/journal.pone.0105189

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