Route of infectious bronchitis virus vaccination determines the type and magnitude of immune responses in table egg laying hens

Abstract

Chicken immune responses to infectious bronchitis virus (IBV) vaccination can depend on route of administration, vaccine strain and bird age. Typically for layer chickens, IBV vaccinations are administered by spray in the hatchery at day-old and boosted at intervals with live vaccines via drinking water (DW). Knowledge of live attenuated IBV vaccine virus kinetics and the immune response in egg-laying hens is exceptionally limited. Here, we demonstrated dissemination of vaccine viruses and differences in hen innate, mucosal, cellular and humoral immune responses following vaccination with Massachusetts or 793B strains, administered by DW or oculonasal (ON) routes. Detection of IBV in the Mass-vaccinated groups was greater during early time-points, however, 793B was detected more frequently at later timepoints. Viral RNA loads in the Harderian gland and turbinate tissues were significantly higher for ON-Mass compared to all other vaccinated groups. Lachrymal fluid IgY levels were significantly greater than the control at 14 days post-vaccination (dpv) for both vaccine serotypes, and IgA mRNA levels were significantly greater in ON-vaccinated groups compared to DW-vaccinated groups, demonstrating robust mucosal immune responses. Cell mediated immune gene transcripts (CD8-α and CD8-β) were up-regulated in turbinate and trachea tissues. For both vaccines, dissemination and vaccine virus clearance was slower when given by DW compared to the ON route. For ON administration, both vaccines induced comparable levels of mucosal immunity. The Mass vaccine induced cellular immunity to similar levels regardless of vaccination method. When given either by ON or DW, 793B vaccination induced significantly higher levels of humoral immunity.

Keywords: Avian coronavirus; cellular and humoral immunity; gene transcription; infectious bronchitis virus; innate; layer chicken; mucosal; vaccination.

Figure 1

Mean anti-IBV ELISA antibody titres of the control (n   = 8) and vaccinated groups ( n  = 5) at 0, 7 and 14 dpv. Data are expressed as mean values  ±  SEM. The positive cut-off titre of 396 is indicated with a dotted line. Different letters indicate significant differences (p  < 0.05) within time points, determined using one-way ANOVA. D0:   Day 0; DW:  Drinking water; ON:  Oculonasal.

Figure 2

Measurement of IBV-specific A IgA and B IgY titres using indirect monoclonal ELISA from lachrymal fluid. Data are expressed as mean of corrected optical density (COD)  ±  SEM. Different letters indicate significant differences (p  < 0.05) within time points, determined using one-way ANOVA.

Figure 3

Relative mRNA expression of IgA in A turbinate and B tracheal tissues, and IgY in C turbinate and D trachea tissues at 1, 3, and 5 dpv. Data are expressed as log₂ mean fold change compared to the control group  ±  SEM. Expression is calculated based on double delta Ct (ΔΔCt) values. Different letters indicate significant differences (p  <  0.05) within time points, determined using one-way ANOVA.

Figure 4

Quantification of viral RNA from OP swabs, expressed as log-relative equivalent units (REUs) of RNA. Samples were taken at 1, 3, 5, 7, and 14 dpv. Data presented as the mean  ±  SEM. Different letters indicate significant (p  < 0.05) differences within groups.

Figure 5

Quantification of viral RNA from CL swabs, expressed as log-relative equivalent units (REUs) of RNA. Samples were taken at 1, 3, 5, 7, and 14 dpv. Data presented as the mean  ±  SEM. Different letters indicate significant (p  < 0.05) differences within groups.

Figure 6

Quantification of viral RNA expressed as a log REU of RNA in the A HG, B turbinate, C choanal cleft, D trachea, E caecal tonsil and F kidney samples. Data presented as the mean  ±  SEM. Different letters indicate significant (p  < 0.05) differences within time points.

Figure 7

Expression profile of host genes in Harderian gland, turbinate, choanal cleft and trachea tissue following IBV Mass or IBV 793B vaccination of layer chickens. Data is shown as the log₂ fold change when compared to the non-vaccinated group. Different letters indicate significant differences (p  < 0.05) in fold change between vaccine strains and vaccination route on the same sample day.

Figure 8

Relative mRNA expression of A CD8-α in turbinate tissue, B CD8-α in trachea tissue, C CD8-β in turbinate tissue and D CD8-β in trachea tissue at 1, 3, and 5 dpv. Data are expressed as log₂ mean fold change compared to the control group  ±  SEM. Expression is calculated based on double delta Ct (ΔΔCt) values. Different letters indicate significant differences (p  < 0.05) within time points, determined using one-way ANOVA.