Effect of Lactobacillus plantarum LP160 with Freeze–Thaw Resistance Characteristics on Fermentation, Bacterial Community, and Metabolomics of Oat Silage in Qinghai–Tibet Plateau

Freeze–thaw cycles on the Qinghai–Tibetan Plateau inhibit microbial activity and challenge silage preservation. This paper aimed to elucidate how an indigenous, freeze–thaw-resistant Lactobacillus plantarum strain (LP160) improves oat silage quality under such stress. Oats were ensiled for 60 days under constant 20 °C (t) or freeze–thaw cycles (12 h at 20 °C/−5 °C; s) with or without LP160 inoculation. Samples after ensiling and 5-day aerobic exposure were analyzed for fermentation parameters, nutrients, microbiome, and non-targeted metabolomics using liquid chromatography–tandem mass spectrometry (LC-MS/MS). LP160 inoculation improved silage quality, as shown by the lower pH, ammoniacal nitrogen, neutral detergent fiber, acid detergent fiber contents as well as the greater amount of lactic acid. Key findings demonstrated that LP160 inoculation significantly enhanced Lactobacillus dominance, effectively curbed the growth of detrimental bacteria like Mucor, and regulated the microbial structure. During the aerobic exposure phase, the microbial community structures and successions varied under different temperature treatments. When inoculated under freeze–thaw conditions, the genus Bacillus increased, while Paenibacillus was not impeded. A total of 943 metabolites were identified, predominantly comprising amino acids, fatty acids, and the like. The expressions of metabolites with antioxidant and antibacterial properties were upregulated with LP160 inoculation. This led to the inhibition of protein hydrolysis and a reduction in ammonia–nitrogen production. The results of correlation analysis indicated that inoculating LP160 suppressed the proliferation of Mucor and enhanced the abundance of Torulaspora; meanwhile, the expression of L-palmitoylcarnitine involved in the fatty acid degradation pathway and fatty acid metabolism pathway was inhibited along with the generation of ammonia–nitrogen. Consequently, the degradation of fatty acids and proteins was restrained. The results of this paper provided new insights into the silage under freeze–thaw conditions.