Chicken IgY Isotype Control

The type VI secretion system (T6SS) has recently emerged as a new pattern of protein secretions in Campylobacter jejuni (C. jejuni). Within the T6SS cluster, hemolysin co-regulated protein (hcp) is considered as a hallmark of functional T6SS and holds key role in bacterial virulence. As poultry is the primary reservoir of C. jejuni and the major sources for human infection, we evaluated the capacity of recombinant hcp (rhcp) immunization in blocking C. jejuni colonization in chickens with an aim to control bacterial transmission to humans via poultry food chain. Considering the mucosal route is the primary portal for C. jejuni entry and gut mucosa offers the apposite site for C. jejuni adherence, we investigated the immune-protective potential of intra-gastric administration of rhcp using chitosan-based nanoparticles. To achieve this goal, full length coding sequence of hcp gene from C. jejuni was cloned and expressed in E. coli. Purified rhcp was entrapped in chitosan-Sodium tripolyphosphate nanoparticles (CS-TPP NPs) and orally gavaged in chickens. Our results suggest that intra-gastric immunization of CS-TPP-rhcp induces consistent and steady increase in intestinal (sIgA) and systemic antibody (IgY) response against rhcp with significant reduction in cecal load of C. jejuni. The protection afforded by rhcp associated cellular responses with Th1 and Th17 profile in terms of increased expression of NFkB, IL-1 beta, IL-8, IL-6, IFN-gamma and IL-17 A genes. Though systemic immunization of rhcp with IFA resulting in a robust systemic (IgY) and local (sIgA) antibody response, mucosal administration of rhcp loaded CS-TPP NPs was found to be superior in terms of bacterial clearance. Altogether, present study suggests that chitosan based infra-gastric delivery of rhcp have several advantages over the injectable composition and could be a promising vaccine approach to effectively control C. jejuni colonization in chickens.

1. Immunological reagents for wild, non-model species are limited or often non-existent for many species. 2. In this study, we compare the reactivity of a newanti-passerine IgY secondary antibody with existing secondary antibodies developed for use with birds. Samples from 41 species from the following six avian orders were analysed: Anseriformes (1 family, 1 species), Columbiformes (1 family, 2 species), Galliformes (1 family, 1 species), Passeriformes (16 families, 34 species), Piciformes (1 family, 2 species) and Suliformes (1 family, 1 species). Direct ELISAs were performed to detect total IgY using goat anti-passerine IgY, goat anti-chicken IgY or goat anti-bird IgY secondary antibodies. 3. The anti-passerine antibody exhibited significantly higher IgY reactivity compared to the anti-chicken and/or anti-bird antibodies in 80% of the passerine families tested. Birds in the order Piciformes (woodpeckers) and order Suliformes (cormorants) were poorly detected by all three secondary antibodies. A comparison of serum and plasma IgY levels was made within the same individuals for two passerine species (house finch and white-crowned sparrow), and serum exhibited significantly more IgY than the plasma for all three secondary antibodies. This result indicates that serummay be preferred to plasma whenmeasuring total antibody levels in blood. 4. This study indicates that the anti-passerine IgY secondary antibody can effectively be used in immunological assays to detect passerine IgY for species in most passerine families and is preferred over anti-chicken and anti-bird secondary antibodies for the majority of passerine species. This anti-passerine antibody will allow for more accurate detection and quantification of IgY in more wild bird species than was possible with previously available secondary antibodies.