P93: Bacterial Population Dynamics and Chemical Transformations During the Ensiling of Medicago sativa (Alfalfa) and Subsequent Exposure to Air

The chemical and bacterial population dynamics that occurred during the process of ensiling alfalfa and subsequent exposure to oxygen were examined using standard chemical techniques and by constructing 16S rRNA gene sequence libraries containing over 1,500 clones each. In the before ensiling library (BE), the vast majority of the bacterial sequences were associated with the phylum Proteobacter (>89%) of which most were related to the family Enterobacteriaceae. After 40 days post-ensiling (PE) the sequences shifted to a predominance of those associated with the phylum Firmicutes (>70%), most of which were associated with the order Lactobacillales; however, the library still contained a substantial amount of sequences related to the phylum Protebacteria (>25%). To simulate an open silage pile, we exposed the silage to 3 L of oxygen per day for 20 days (PO) which selected for greater percentages of sequences related to the phylum Firmicutes (>95%). The levels of diversity within the BE and the PE libraries were similar in richness, with 157 and 167 OTU, respectively. However the PO library contained only 83 OTU. The Shannon indexes reflected these values with 2.63, 3.22 and 1.93 for the BE, PE and PO libraries, respectively. Chemically the concentrations of soluble protein, crude fat and total volatile fatty acids all increased; while the concentrations of ammonia, sugar, starch, and pH were reduced. The total weight of the silage decreased by >10%. The weight loss was concomitant with emissions of volatile gas containing, in decreasing order: CO2, ethanol, methanol, ammonia, and oxides of nitrogen.