Blood proteome profiling for biomarker discovery in broilers with necrotic enteritis.

Analysis of the blood proteome allows identification of proteins related to changes upon certain physiological conditions. The pathophysiology of necrotic enteritis (NE) has been extensively studied. While intestinal changes have been very well documented, data addressing NE-induced alterations in the blood proteome are scant, although these might have merit in diagnostics. In light of recent technological advancements in proteomics and pressing need for tools to access gut health, the current study employs mass-spectrometry (MS)-based proteomics to identify biomarkers for gastrointestinal health of chickens. Here, we report findings of an untargeted proteomics investigation conducted on blood plasma in chickens under NE challenge. Two MS-strategies were used for analysis: conventional data dependent acquisition coupled to standard nanoflow liquid chromatography (LC) (nano-DDA) and recently-developed data independent acquisition coupled to an Evosep One LC system (Evo-DIA). Despite superior completeness and quantification of the Evo-DIA-acquired data, high degree of agreement in identification and quantification was observed between both approaches. Additionally, we identified 15 differentially expressed proteins (shared by nano-DDA and Evo-DIA) that represent responses of animals to infection and may serve as potential biomarkers. Experimental validation through ELISA immunoassays and targeted MS for selected regulated proteins (CFD, HPS5, and MASP2) confirmed medium-to-high levels of inter-protein correlation. A GSEA analysis revealed enrichment in a number of processes related to adaptive and humoral immunity, immune activation and response in infected animals. Data are available via ProteomeXchange with identifiers PXD050461, PXD050473, and PXD061607.