Pre-fermented juices for clean and efficient production of mulberry silage: Insights from the microbiome, metabolome and in vitro rumen fermentation

Mulberry ( Morus alba L.) is a new bioresource for alleviating the feed supply crisis. This study evaluated pre-fermented mulberry juice (M), pre-fermented red clover juice (R), and pre-fermented grape pomace juice (G) to achieve clean and efficient production of mulberry silage. R treatment enriched Enterococcus (LDA > 5) and up-regulated L-aspartic acid and fumaric acid; M treatment enriched Lactobacillus (LDA > 5) and up-regulated colchicine and 3,4-dihydroxybenzaldehyde; and G treatment enriched Kosakonia (LDA > 4) and Lactiplantibacillus (LDA > 5) and up-regulated p -coumaric acid and chlorogenic acid. Pre-fermented juices modulated the microbiota community and metabolites of mulberry silage, thus regulating silage quality, especially in dry matter, crude protein, total nitrogen, nonprotein nitrogen, ammonia nitrogen, pH, lactic acid, and acetic acid. Furthermore, pre-fermented juices improved the in vitro rumen fermentation. R and M treatments increased in vitro gas production while reducing CO 2 emissions without increasing CH 4 production, illustrating that R and M treatments alleviated greenhouse gas production. Pre-fermented juices promoted in vitro rumen fermentation and enhanced the nutrition and fermentation quality of mulberry silages. This study revealed that pre-fermented juices achieved clean and efficient production of mulberry silage, which may promote sustainable development of the livestock industry. • Pre-fermented juices modulated microbiota community and metabolic pathways in different ways. • R and M treatments increased in vitro gas production while reducing CO 2 emissions. • Bioactive metabolites were up-regulated by pre-fermented juices.