An attenuated duck plague virus (DPV) vaccine induces both systemic and mucosal immune responses to protect ducks against virulent DPV infection

Abstract

Duck plague (DP) is a severe disease caused by DP virus (DPV). Control of the disease is recognized as one of the biggest challenges in avian medicine. Vaccination is an efficient way to control DPV, and an attenuated vaccine is the main routine vaccine. The attenuated DPV vaccine strain CHa is a modified live vaccine, but the systemic and mucosal immune responses induced by this vaccine have been poorly understood. In this study, the immunogenicity and efficacy of the vaccine were evaluated after subcutaneous immunization of ducks. CD4(+) and CD8(+) T cells were counted by flow cytometry, and humoral and mucosal Ig antibodies were analyzed by enzyme-linked immunosorbent assay (ELISA). The results showed that high levels of T cells and Ig antibodies were present postimmunization and that there were more CD4(+) T cells than CD8(+) T cells. Titers of humoral IgG were higher than those of humoral IgA. Local IgA was found in each sample, whereas local IgG was found only in the spleen, thymus, bursa of Fabricius, harderian gland, liver, bile, and lung. In a protection assay, the attenuated DPV vaccine completely protected ducks against 1,000 50% lethal doses (LD50) of the lethal DPV strain CHv via oral infection. These data suggest that this subcutaneous vaccine elicits sufficient systemic and mucosal immune responses against lethal DPV challenge to be protective in ducks. This study provides broad insights into understanding the immune responses to the attenuated DPV vaccine strain CHa through subcutaneous immunization in ducks.

FIG 1

T lymphocytes in peripheral blood following subcutaneous immunization with the attenuated DPV vaccine strain CHa. (A) CD4⁺ T lymphocytes in peripheral blood were collected at 1, 2, 3, 4, 6, and 8 weeks and analyzed by flow cytometry. (B) CD8⁺ T lymphocytes in peripheral blood were collected at 1, 2, 3, 4, 6, and 8 weeks and analyzed by flow cytometry. The data presented are the mean values for three samples at each time point. *, P < 0.05 compared to control group; **, P < 0.01 compared to control group; ***, P < 0.001 compared to control group. PBLCs, peripheral blood lymphocytes.

FIG 2

DPV-specific IgA and IgG in sera were detected by antigen-capture ELISA. Sera were collected at 1, 2, 3, 4, 5, 6, and 8 weeks, and the level of IgA (A) or IgG (B) in each sample was detected and compared with that for the negative-control group. *, P < 0.05 compared to control group; **, P < 0.01 compared to control group.

FIG 3

Mucosal IgA responses were assessed by ELISA. Spleens, bursas of Fabricius, thymuses, harderian glands, lungs, tracheas, livers, bile, duodena, jejuna, ilea, ceca, and rectums were obtained from the vaccinated group. IgA responses in immune (A), digestive (B), and respiratory (C) organs were tested by DPV antigen-capture ELISA.

FIG 4

DPV-specific mucosal IgG responses were tested by ELISA. Spleens, thymuses, harderian glands, bursas of Fabricius, livers, bile, and lungs were obtained from the vaccinated group at 1, 2, 3, 4, 5, 6, and 8 weeks. The specific IgG in different organs was detected by DPV antigen-capture ELISA.

FIG 5

Survival curve after challenge with lethal DPV strain CHv. Ten ducks from the vaccinated group or negative-control group were orally challenged with 1,000 LD₅₀ of DPV strain CHv on the 6th week after the first immunization. Mortality was monitored daily for 10 days after challenge. The figure was drawn as a Kaplan-Meier survival curve. The y axis shows the cumulative (Cum) survival calculated by the Kaplan-Meier method (P < 0.01).