Mapping B-cell responses to Salmonella enterica serovars Typhimurium and Enteritidis in chickens for the discrimination of infected from vaccinated animals

Abstract

Serological surveillance and vaccination are important strategies for controlling infectious diseases of food production animals. However, the compatibility of these strategies is limited by a lack of assays capable of differentiating infected from vaccinated animals (DIVA tests) for established killed or attenuated vaccines. Here, we used next generation phage-display (NGPD) and a 2-proportion Z score analysis to identify peptides that were preferentially bound by IgY from chickens infected with Salmonella Typhimurium or S. Enteritidis compared to IgY from vaccinates, for both an attenuated and an inactivated commercial vaccine. Peptides that were highly enriched against IgY from at least 4 out of 10 infected chickens were selected: 18 and 12 peptides for the killed and attenuated vaccines, respectively. The ten most discriminatory peptides for each vaccine were identified in an ELISA using a training set of IgY samples. These peptides were then used in multi-peptide assays that, when analysing a wider set of samples from infected and vaccinated animals, diagnosed infection with 100% sensitivity and specificity. The data describes a method for the development of DIVA assays for conventional attenuated and killed vaccines.

Figure 1. IgY recognition of Salmonella epitopes/mimotopes.

Purified IgY from infected (S. Typhimurium or S. Enteritidis, n = 16 and n = 19, respectively, numbers 1–16 and 17–35 respectively in (C,D) and vaccinated (n = 20 for both the killed (A,C) and attenuated (B,D) vaccines, numbers 1–20 in (C,D) chickens was analysed for binding to synthetic peptides. Bound IgY was detected with an anti-IgY-AP conjugate. All samples were analysed in duplicate. Example discriminatory individual peptides for the killed (A) or attenuated (B) vaccine are shown. The cut-off values (dotted lines in (A,B) were calculated as the mean +5SD of the signals for the 10 vaccinates used in the panning steps (training cohort, this is also applied for analysis in (C,D). The 10 most discriminatory peptides are shown as a multi-peptide ELISA for the killed (C) and attenuated (D) vaccine where binding of an IgY sample above the cut-off value for a peptide is shown as a grey box.