Abstract Background Elucidating the mechanism of fiber transformation underlying microbial metabolism is critical for improving fiber-rich silage digestibility and preserving silage energy for ruminant nutrient absorption. However, few studies have combined quantitative microbial function and transformation products in silage to explain this mechanism. Here, we constructed a workflow to detect the substrates and products of fiber transformation in mixed silage of Sesbania cannabina and sweet sorghum (SS) and combined the absolute quantification 16S rRNA sequencing to reveal this mechanism. Results The synergistic effect of Lactobacillus cocktail and cellulase (LC) simplified the microbial diversity and minimized the microbial quantity, making Lentilactobacillus buchneri the dominant species in SS silage. As a result, the LC-treated silage had greater lactic acid content, lower pH value, and less NH3-N content. The indigestible fibers were significantly decreased due to the synergistic effect of the Lactobacillus cocktail and cellulase. Changes in microbial structure during ensiling also resulted in metabolic alterations. The increased levels of microbial enzymes, including β-glucosidase and sucrose phosphorylase, involved in starch and sucrose metabolism led to the enrichment of monosaccharides (including glucose, xylose, mannose, galactose, ribose, rhamnose, and arabinose) in the LC-treated silage. We found that L. buchneri was positively associated with β-glucosidase and sucrose phosphorylase, reflecting the crucial contribution of L. buchneri to fiber decomposition in SS silage. Conclusion Using an absolute quantitative microbiome, we found that LC treatment decreased the microbial biomass in SS silage, which in turn promoted the energy preservation in the SS silage. The cooperative interaction of the Lactobacillus cocktail and cellulase improved the fiber decomposition and in vitro dry matter digestibility rate by changing the microbiome structure and function in the SS silage, providing guidance and support for future fiber-rich silage production in the saline-alkaline region. Graphical Abstract
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