New Insights Into Dietary L-Glutamate and L-Aspartate Modulation of Hematology, Immune Responses, and Metabolite Profiles in Enterotoxigenic Escherichia coli Challenged Piglets

Background/Objectives: L-glutamate (Glu) and L-aspartate (Asp) are key intermediates in nitrogen metabolism and tricarboxylic acid cycle activity, linking intestinal energy metabolism with immune function. This study investigated how dietary Glu and Asp supplementation modulates immune responses and metabolic reprogramming in weaned pigs challenged with F18 enterotoxigenic Escherichia coli (ETEC). Methods: Forty-nine piglets (24 d old; 8.18 ± 1.54 kg body weight) were randomly assigned to seven treatments (n = 7/treatment): unchallenged control (NC), ETEC-challenged control (PC), 1% or 2% Glu, 1% or 2% Asp, and an antibiotic control. The experiment was conducted from d −7 to d 14 post-inoculation (PI). Hematological indices, serum biomarkers, intestinal cytokine gene expression, and untargeted metabolomic profiling of serum, ileal mucosa, and ileal digesta were evaluated. Results: On day 2 PI, 1% Glu reduced the neutrophil-to-lymphocyte ratio, whereas 2% Asp showed an elevated ratio. Supplementation of 1% Asp increased serum total protein on d 2 and d 5 PI. On d 14 PI, 1% Glu enhanced jejunal IL-17A and IL-22 expression, while 2% Asp reduced jejunal IL-6 expression compared with PC. Ileal IL-12 expression increased with 1% Glu and 2% Asp, whereas jejunal IL-12 expression decreased with 2% Glu and 2% Asp. Untargeted metabolomics revealed distinct treatment-dependent separations. Differential metabolite profiling and pathway enrichment analyses demonstrated coordinated alterations in amino acid metabolism, purine metabolism, lipid metabolism, and energy-related pathways across serum and intestinal compartments. Conclusions: Collectively, Glu and Asp supplementation reshaped host metabolic networks during ETEC challenge, indicating their roles in modulating metabolic adaptation and intestinal immune–metabolic crosstalk under enteric stress.