Community Synergy of Lactic Acid Bacteria and Cleaner Fermentation of Oat Silage Prepared with a Multispecies Microbial Inoculant

Abstract

To investigate community synergy of lactic acid bacteria (LAB) and cleaner fermentation of oat silage, oat silages were prepared with or without (control) commercial LAB inoculants LI1 (containing Lactiplantibacillus plantarum, Lentilactobacillus buchneri, Lacticaseibacillus paracasei, and Pediococcus acidilactici) and LI2 (containing Lactiplantibacillus plantarum and Lentilactobacillus buchneri). The microbial community, LAB synergy, and cleaner fermentation were analyzed at 1, 3, 6, 15, 35, and 90 days of ensiling. The LAB inoculant improved fermentation quality, with significantly (P < 0.05) lower pH, ammonia nitrogen content, and gas production and higher lactic acid and acetic acid contents than those of the control. Enterobacteriaceae was the main bacterial community in early stage of fermentation, which utilizes sugar to produce CO₂ gas, causing dry matter (DM) and energy loss. As fermentation progressed, the microbial diversity decreased, and the microbial community shifted from Gram-negative to Gram-positive bacteria. The inoculation of multispecies LAB displayed community synergy; Pediococcus acidilactici formed a dominant community in the early stage of fermentation, which produced an acid and anaerobic environment for the subsequent growth of Lentilactobacillus and Lacticaseibacillus species, thus forming a LAB-dominated microbial community. The predicted functional profile indicated that the silage inoculated with LI1 enhanced the carbohydrate metabolism pathway but inhibited the amino acid metabolism pathway, which played a role in promoting faster lactic acid production, reducing the decomposition of protein to ammonia nitrogen, and improving the fermentation quality of silage. Therefore, oat silage can be processed to high-quality and cleaner fermented feed by using an LAB inoculant, and LI1 showed better efficiency than LI2. IMPORTANCE Oat natural silage is rich in Enterobacteriaceae, increasing gas production and fermentation loss. Lactic acid bacteria interact synergistically to form a dominant community during ensiling. Pediococci grow vigorously in the early stage of fermentation and create an anaerobic environment. Lactobacilli inhibit the harmful microorganisms and result in cleaner fermentation of oat silage.

Keywords: clean fermentation; community synergy; gas production; lactic acid bacteria; oat.

FIG 1

Relative abundance of bacteria community in oat and silage at the phylum (a) and genus (b) levels across different groups and fermentation times. LI1, silage inoculated with Lactiplantibacillus plantarum, Lacticaseibacillus casei, Lentilactobacillus buchneri, and Pediococcus acidilactici; LI2, silage inoculated with Lactiplantibacillus plantarum and Lentilactobacillus buchneri; D1, D3, D6, D15, D35, and D90, days 1, 3, 6, 15, 35, and 90 of silage, respectively.

FIG 2

Heat map of prominent bacterial genera (20 most abundant genera) for oat silage before and after ensiling for 1, 3, 6, 15, 35, and 90 days with or without LI1 and LI2.

FIG 3

One-way analysis of variance bar plots of the 10 most abundant genera among different oat silage treatments. *, 0.01 < P ≤ 0.05; **, 0.001 < P ≤ 0.01; ***, P ≤ 0.001.

FIG 4

Association analysis between bacterial genera and quality variables. Quality variables are displayed horizontally, and the bacterial genera are displayed vertically. The corresponding value of the middle heat map is the Pearson correlation coefficient r, which ranges between −1 and 1; an r value of <0 indicates a negative correlation (blue), and a value of >0 indicates a positive correlation (red). *, P < 0.05; **, P < 0.01; ***, P < 0.001. (a) Correlations between bacterial genera and quality variables in different silage groups. (b) Correlations between bacterial genera and quality variables at different ensiling time points (1, 3, 6, 15, 35 and 90 days of ensiling). LBC, lactate buffer capacity; WSC, water-soluble carbohydrate; LA, lactic acid; AA, acetic acid; AN, ammonia nitrogen; GP, gas production.

FIG 5

Level 2 KEGG orthologous sequences of ensiled oat as influenced by additives and ensiling time. *, 0.01 < P ≤ 0.05; **, 0.001 < P ≤ 0.01; ***, P ≤ 0.001. Functional prediction of bacterial changes in oat after fermentation was made using PICRUSt2.