Identification and isolation of Lactobacillus fructivorans from wilted alfalfa silage with and without molasses

Aim: To gain insights into the bacterial species associated with anaerobic storage and aerobic stability of alfalfa silage. Methods and Results: Wilted alfalfa silage (498 g dry matter kg^-1) was prepared with and without the addition of molasses. Aerobic spoilage tests were conducted at 5, 10 and 60 days after ensiling. The composition of fermentation products and the bacterial communities of silage were determined at 1, 3, 5 and 7 days after silo opening. Silage without molasses had small amounts of lactic and acetic acids detectable at silo opening but resisted deterioration due to aerobic spoilage for at least 5 days after opening. Resistance to aerobic deterioration in silage increased with the addition of molasses. The predominant bacterial species in molasses-added silage was Lactobacillus fructivorans, which was detected by denaturing gradient gel electrophoresis analysis. Different bacterial growth media were used for Lact. fructivorans isolation from alfalfa silage with added molasses: isolation was successful using liver infusion sake medium, but was unsuccessful when de Man, Rogosa and Sharpe medium was used. Conclusion: A nonconventional lactic acid bacterium (LAB) species may be involved in the high aerobic stability of alfalfa silage. Significance and Impact of the Study: The findings demonstrate that culture-independent microbiota analysis may be useful in the isolation and identification of nonconventional LAB species involved in fermentation and the aerobic stability of silage.