Effects of additive on the ensilage quality and anaerobic digestion performance of hybrid Pennisetum

Hybrid Pennisetum is one species of the perennial forage grass in the genus Pennisetum of the Poaceae family. It is also recognized by its exceptional adaptability, including drought tolerance, salinity resistance, waterlogging resilience, and ability to thrive in nutrient-poor soils. Hybrid Pennisetum has been widely utilized in animal feed production for the manufacture and renewable energy. A highly promising feedstock can also provide for anaerobic digestion, with a methane yield of 104-328 mL/g. However, a critical challenge has hindered its year-round utilization, due to the inherent conflict between seasonal harvesting cycles and the demand for continuous biomass supply. Consequently, it is often required for the long-term preservation of hybrid Pennisetum biomass for its sustainable and efficient application. Among them, some ensilage has been widely adopted as a mainstream preservation, due to its operational simplicity, space efficiency, as well as the nutritional integrity and biomass quality of forage materials. Specifically, the silage is essentially a microbial fermentation. The exogenous agents can be added to effectively regulate the fermentation for the product quality. Previous studies have confirmed that the additives have a synergistic effect on the ensilage and anaerobic digestion. However, there are still scientific questions about the variations in the microbial community structure caused by additives during ensilage. This study aims to explore the metabolic pathways of the microorganisms in subsequent anaerobic digestion, in order to improve the ensilage quality and methane performance of hybrid Pennisetum. A systematic investigation was also made to clarify the effects of the lactic acid bacteria and distiller's grains additives on the ensilage and anaerobic digestion. High-throughput sequencing was used to determine the variation in the microbial communities during ensilage and anaerobic digestion. The metabolic network of the key microorganisms was obtained for the synergistic mechanism and regulatory pathways. Five experimental groups were set, including the fresh grass group (C), control group (CK), lactic acid bacteria additive group (A), distiller's grains additive group (B), as well as the composite additive group of lactic acid bacteria and distiller's grains (AB). The quality of hybrid Pennisetum was systematically examined in the 90-day ensilage pretreatment period. The methane production performance of ensilage hybrid Pennisetum was also evaluated in the 30-day anaerobic digestion period. The results showed that compared with the group without additives (CK) in the ensilage, the lactic acid bacteria and vinasse composite additive reduced the pH value and total ammonia nitrogen of ensilage samples to 4.77 and 67.6 mg/L, respectively, whereas there was an increase in the concentration of acetic acid by 8%. The relative abundance of the beneficial microorganisms increased by 7% to 231%, such as Lactobacillus in ensilage. In the anaerobic digestion stage, the cumulative methane yield of the ensilage hybrid Pennisetum with the lactic acid bacteria and vinasse composite additives reached 322.97 mL/g, which was 65% higher than that of the fresh grass group. Clostridium sensu sirico_1, Terrisporobacter, Methanosarcina, and Methanobrevibacter were dominant microorganisms in anaerobic digestion. Acetoclastic and hydrogenotrophic forms were the main pathways for methane production. Overall, the combination of the lactic acid bacteria and vinasse as additives can synergistically improve the ensilage quality and anaerobic digestion of Hybrid Pennisetum. The efficient storage of green raw materials, such as the hybrid Pennisetum, can be obtained in order to improve the energy conversion efficiency of the raw materials. It is of great significance to support the national "dual carbon" goal in renewable energy.