Microbial and metabolic responses of maize silage to leaf blight: implications for fermentation and protein stability

Foliar fungal diseases lead to a significant reduction in dry matter of plants, thereby negatively affecting silage fermentation kinetics. This study investigated the effects of leaf blight on ensiling kinetics, microbial succession, and nitrogen metabolism in whole-crop maize, and further examined whether inoculation with Lactiplantibacillus plantarum HT1 could mitigate disease-induced proteolysis through remodeling of the metabolic pathways. Three treatments were established: (i) healthy maize silage (CON), (ii) maize silage affected by leaf blight (DCON), and (iii) maize silage affected by leaf blight and inoculated with L. plantarum HT1 (HT1, 1 × 10⁵ cfu/g FM based on the fresh weight basis). Samples of 300 g fresh material were packed into polyethylene vacuum bags (300 × 400 × 0.2 mm) and vacuum-sealed to establish anaerobic conditions for 60 days of ensiling. Before ensiling, DCON had significantly lower crude protein content (7.06% vs. 8.91% DM, P = 0.001) compared with CON. In addition, the WSC content markedly decreased by leaf blight (88.7 vs. 119 g/kg DM, P = 0.016). LAB abundance was significantly lower (3.22 vs. 4.22 log10 CFU/g FM, P = 0.022), whereas mold counts sharply increased (4.25 vs. 3.22 log10 CFU/g FM, P = 0.001) in DCON than in CON. The DCON showed elevated pH, significantly reduced lactic acid content, and markedly increased butyric acid content compared with those of the CON (P < 0.05). DCON had the highest NH₃-N content (17.2 g/kg TN), exceeding those of CON (12.3 g/kg TN) and HT1 (10.3 g/kg TN, P = 0.005). Aminopeptidase and carboxypeptidase activities increased to 46.4 and 167 U·h^{- 1}·g FM^{- 1}, respectively, which were much higher than those of the CON (27.4 and 140 U·h^{- 1}·g FM^{- 1}). Microbiome β-diversity separated DCON from CON and HT1, with enrichment of putative proteolytic taxa. Metabolomics indicated upregulated amino acid degradation (branched chain and glutamate pathways) and disrupted nitrogen homeostasis in the DCON treatment. Leaf blight created an early high-pH window that amplified proteolysis and nitrogen loss through coordinated shifts in substrates, microbiota, and amino acid catabolism. Inoculation with L. plantarum HT1 caused rapid acidification, curtailed proteolysis, and rewired metabolic and community networks toward a healthy state, achieving coordinated restoration of fermentation quality and protein preservation in disease-challenged maize.