Contained Ensiling of High-Lipid Perennial Ryegrass: Fermentation Quality, Fatty Acid Retention, and Storage Stability

Genetically modified (GM) forage crops engineered to accumulate elevated levels of lipids offer potential benefits for ruminant nutrition and greenhouse gas mitigation. However, robust and reproducible workflows for producing, harvesting, and preserving GM forage biomass under containment remain a critical bottleneck, particularly where regulatory constraints preclude field-scale evaluation. Here, we describe a controlled-environment workflow for the repeated cultivation, harvesting, and ensiling of GM high-metabolizable-energy (HME) perennial ryegrass and corresponding null controls. Plants were grown under greenhouse containment, subjected to multiple regrowth cycles, and harvested biomass was wilted and ensiled using small-scale laboratory silos. Silage fermentation characteristics, total lipid content, and fatty acid (FA) composition were assessed following short- and long-term storage. Over 16 months, approximately 130 kg dry matter (DM) of each genotype was produced across multiple harvests and ensiling batches. Seasonal variation strongly influenced herbage composition, with water-soluble carbohydrate concentrations 4–5-fold higher in spring–summer than autumn–winter. Following ensiling, HME silage consistently retained elevated FA content compared with null controls (4.85% vs. 2.75% DM) and higher gross energy (18.1 vs. 17.5 MJ kg−1 DM). FA profiling indicated that major FA classes in HME were preserved across storage durations. After 342 days of storage, HME silage maintained 76% higher FA content, 4% greater DM digestibility, and 0.3–0.8 MJ kg−1 DM higher metabolizable energy. Both genotypes exhibited good fermentation quality, with pH consistently below 4.1 and adequate lactic acid production. This study does not evaluate animal performance or methane mitigation outcomes but establishes a practical and reproducible methodology for generating characterized GM silage material under containment suitable for subsequent in vivo studies, addressing a key translational gap between GM forage development and animal-based evaluation.