The Microbiota Dynamics of Alfalfa Silage During Ensiling and After Air Exposure, and the Metabolomics After Air Exposure Are Affected by Lactobacillus casei and Cellulase Addition

Abstract

Both inoculants treatment and enzyme treatment promote the reproduction of lactic acid bacteria (LAB) to produce enough lactic acid to lower pH in silage. The present study investigated the microbial community and metabolome in cellulase, Lactobacillus casei, and air treated alfalfa silage. Chopped and wilted alfalfa (first cutting, 29% dry matter) was ensiled without (CON) or with L. casei (1 × 106 cfu g–1 fresh matter) (LC) or cellulase (20,000IU, 0.5% of fresh matter) (CE) for 56 days, then exposed to air for 3 days (PO). Greater ensiling quality was observed in LC and CE, which had lower pH and higher lactic acid content than CON at 56 days of ensiling and 3 days post-oxygen exposure. Air exposure was associated with decreased lactic acid concentrations and increased yeast and mold counts in all silages. SEM showed that the structure of leaf epicuticular wax crystals were intact in fresh alfalfa, totally decomposed in CON silage, and partly preserved in CE and LC silage. Gas chromatography mass spectrometry revealed that 196 metabolites and 95 differential concentration were present in the 3 days air exposure samples. Most of these metabolites, mainly organic acids, polyols, ketones, aldehydes, are capable of antimicrobial activity. The bacterial communities were obviously different among groups and Lactobacillus developed to a dominant status in all silages. Lactobacillus became dominant in bacterial communities of LC and CE silages from days 7 to 56, and their relative abundances reached 94.17–83.93% at day 56, respectively. For CON silage, until day 56, Lactobacillus dominated the bacterial community with abundance of 75.10%. After 3 days of oxygen exposure, Lactobacillus and Enterococcus were predominant in CON, and Lactobacillus remained dominant in LC and CE silages. The results indicated that, compared to untreated silages, L. casei could be a priority inoculant for alfalfa silage to boost Lactobacillus abundance and improve fermentation quality. Our high-throughput sequencing and gas chromatography mass spectrometry results provide a deep insight into the bacterial community and metabolites in alfalfa silage.

Modulation of Fermentation Quality and Metabolome in Co-ensiling of Sesbania cannabina and Sweet Sorghum by Lactic Acid Bacterial Inoculants

“…Lactose can be utilized by LAB to produce acids and flavor, but it cannot be utilized by pathogenic or spoilage organisms ( Vos and Vaughan, 2010 ). We found that LAB inoculation resulted in accumulated α-lactose, which is beneficial for reducing pH and inhibiting the growth of pathogens during ensiling, according to previous report ( Hu et al, 2020 ). In contrast, organic nitrogen compounds, like histamine with toxicological characteristics, were downregulated with LAB treatment.…”

Influence of Condensed and Hydrolysable Tannins on the Bacterial Community, Protein Degradation, and Fermentation Quality of Alfalfa Silage

“…Hu et al [ 30 ] observed that Proteobacteria were the preponderant bacteria in alfalfa before ensiling, and there was a tendency towards a shift from Proteobacteria to Firmicutes during ensiling. In this study, the addition of tannins increased the relative abundance of Proteobacteria and decreased the relative abundance of Firmicutes in alfalfa silage.…”

Additives Altered Bacterial Communities and Metabolic Profiles in Silage Hybrid Pennisetum

“…In the present study, diosmetin was positively related to Thermoactinomyces daqus and Brevibacillus thermoruber , suggesting that these two species might be involved in the production of diosmetin and other bioactive compounds. Several studies proved that different additives in silage can cause changes in bioactive metabolites, such as polyphenols, flavonoids, and terpenoids ( Guan et al, 2020 ; Hu et al, 2020 ; Xu et al, 2021 ). However, these active substances are rarely used directly as silage additives.…”

“…With the process of fermentation, the activities of microorganisms produce a large number of metabolites, including amino acids, aromatic compounds, fatty acids, flavoring agents, oligosaccharides, peptides, and vitamins ( Guo et al, 2018 ). In addition to lactic acid (LA), acetic acid, propionic acid, and other conventionally detected substances, other metabolites, such as sorbic acid, isovaleric acid, 3-phenyllactic acid, and hydroferulic acid, play important roles in maintaining aerobic stability and improving the fermentation quality of silage ( Hu et al, 2020 ). Metabolomics focuses on all small-molecule metabolites with molecular weights less than 1,000 Da and can monitor the changes in metabolic profiles caused by different treatments.…”