ناهید آقامحمدی

Corn silage is a major source of forage for ruminants, which provides a higher energy level compared to other forages. But it is susceptible to aerobic deterioration, because yeasts utilize the lactic acid, produced by lactic acid bacteria, as a source of energy. Thus, silages become a favor environment for the growth of mold and bacteria. The susceptibility of a silage to the aerobic deterioration is an important factor since it is directly related to its quality and nutritive value. When silage is exposed to air, increase of temperature and pH may occur, resulting in soluble carbohydrates loss and fermentation of the products of microbial metabolism, reducing its quality. Aerobic stability is an important feature in the evaluation of corn silage quality. Researchers reports that the use of lactic acid bacteria can improve fermentation and aerobic stability of silage. This study aimed to investigate the changes in the chemical compositions and concentration of fermentation end products that occur in corn silages with or without microbial inoculants during aerobic exposure and to select bacterial strains that can improve the aerobic stability after silo opening. This study was conducted to select lactic acid bacteria (LAB) strains isolated from corn silage, the intestinal contents of broilers, laying hens, Turkey, Ostrich and assess their effects on the quality and aerobic stability of maize silage. The LAB strains were inoculated into aqueous extract obtained from maize to evaluate their production of metabolites, pH reduction and antimicrobial activity. One hundred and twenty-one strains were isolated from various sources in the Laboratory, which all isolates were considered to be LAB as determined by Gram-stain appearance, catalase test and lactic acid productivity. Analysis of the 16S rDNA sequence of representative strains was used to confirm the presence of the predominant groups. The sequences from the various LAB isolates showed high degrees of similarity to those of the GenBank type strains between 99% and 100%. Five LAB strains that showed the best results were assessed in experimental silos. All treatments were ensiled in laboratory-scale silos for 105 days, and then subjected to an aerobic stability test for 8 days. Silage samples were collected at 0, 4 and 8 days after aerobic exposure to determinate the changes in the chemical compositions, products of fermentation, and to evaluate the silages microbial changes to determine the aerobic deterioration. Data in the experimental design, after opening the silos, were analyzed in a completely randomized design with nine replicates by GLM procedure of SAS software. In aerobic conditions, the data were analyzed as repeated measures in time. The data were analyzed in a completely randomized design by GLM procedure of SAS software. Variance analysis and multiple comparisons of data were performed using the GLM procedure of and the means were separated by Tukey’s test. The results indicated that after 105 days of conservation, all silages had good fermentation characteristics with low pH (<3.80) and ethanol concentration and high lactic acid contents (P<0.01). A linear decrease in pH values and water-soluble carbohydrates contents were observed. Addition of lactic acid bacteria, increased acetic acid, but decreased ethanol contents of inoculated silages compared to the control (P<0.01).  Across treatments, there were significant differences in the yeast populations (P<0.01). The population of yeasts which was initially high in the fresh forage (5.57 log cfu g−1 forage) before ensiling and a drastic reduction in the numbers of yeasts was observed in all silages throughout the experiment. The addition Lactobacillus Fermentum resulted in a higher concentration of acetic acid and reduced populations of yeasts in silage compared to the other silage treatments, and a lower ethanol concentration in the silage (P<0.01). The strains tested were able to modify the fermentative and chemical parameters and the populations of yeasts of silage after aerobic exposure.  Aerobic stability of corn silages was associated with high acetic acid and reduction the populations of yeasts. all Lactobacillus strains promoted an increase in aerobic stability of silage. Addition of lactic acid bacteria, further improved silage aerobic stability with more acetic acid production and reduction the populations of yeasts (P<0.01). After 8 days of aerobic exposure, inoculated silage with Lactobacillus Fermentum remained stable, but the control silage deteriorated as indicated by a reduction in lactic acid and an increase in pH, and numbers of yeast (P<0.01). These results proved the advantage of microbial inoculation. The best Lactobacillus strains is Lactobacillus Fermentum because it provides more stable pH, production higher acetic acid, higher reduction of yeasts and filamentous fungi in maize silage, thereby decreasing the aerobic deterioration by these microorganisms. The results showed that application of inoculants is recommended because it promoted higher production of acetic acid, reducing the populations of yeast and filamentous fungi, a more stable pH and, therefore, improving the aerobic stability of silages. The best inoculation is Lactobacillus Fermentum because it provides higher reduction of yeast and filamentous fungi in maize silage, thereby decreasing the aerobic deterioration by these microorganisms. The Lactobacillus Fermentum strain show the best results in relation to silage aerobic stability.

The effects of lactic acid bacteria as a microbial additive on the fermentation properties of silage are mainly due to the production of beneficial metabolites that can inhibit the growth of pathogenic and destructive microorganisms. Therefore, the ability of a strain to use numerous substrates in forage plants and produced different metabolites can be useful in competition with other microorganisms. This ability can be used as a principle for selecting inoculants. Nevertheless, research has shown that published articles on inoculant selection to limit the growth of these destructive and pathogenic microorganisms are still scarce. Some researchers have reported the effects of microbial inoculants on silage fermentation. However, despite numerous articles on the effects of inoculants on silage fermentation, there is little descriptions for the systematic selection of strains. The aim of this study was to isolate, identify and select strains of lactic acid bacteria with the ability to improve fermentation properties and prevent the growth of pathogenic and destructive microorganisms and evaluate the effect of inoculation of these strains on nutritional value and aerobic stability of maize silages.

This study was performed to select the lactic acid bacterial (LAB) strains isolated from different sources and evaluate their effect on the quality of maize silage. Lactic acid bacteria strains were inoculated to evaluate metabolite production and pH reduction in aqueous extract obtained from maize forage. One hundred and twenty-one strains were isolated from various sources in the laboratory. All isolated strains were gram-positive, catalase-negative and mainly lactic acid-producing, which were considered lactic acid bacteria. 16S ribosomal DNA sequence analysis of 22 representative strains was used to confirm the presence of dominant groups. The sequences of different isolates of lactic acid bacteria had a high degree of similarity to the GeneBank type strains, which showed between 99 and 100% similarity. Four strains of lactic acid bacteria that showed the best results were evaluated in experimental silos. This experiment was performed in a completely randomized design with five treatments (four strains of lactic acid bacteria and one control without inoculum) and nine replications. Inoculants were used in 106 colony forming unit/g of fresh forage to chopped maize forage, which were then ensiled for 105 days. After opening the silos, silages were sampled for analysis of chemical compositions and fermentation products.

Inoculation of lactic acid bacterial strains affected the chemical composition, fermentation properties and microbial population. Inoculation of lactic acid bacterial strains significantly affected the chemical composition and concentration of lactic and acetic acids, ethanol and 1,2-propandiol of silage. Among all treatments, silages inoculated with Lactobacillus fermentum had higher acetate concentration (p < 0.0001), but lactate concentration (p < 0.008) and yeast counts (p < 0.0001) were lower than other silages. Also, inoculation of silage with L. fermentum (strain 16) resulted in silage with higher lactic acid bacterial population and improved aerobic stability after air exposure (p < 0.0001). Silages inoculated with L. fermentum and fallowed by with L. salivarius contained the lowest number of yeasts (p < 0.0001) and filamentous fungi during fermentation. Between silages, silage inoculated with L. fermentum showed higher aerobic stability (p < 0.0001).

The pre-selection method based on the production of metabolites was efficient in selecting new inoculation strains along good correlation with experiments in laboratory silos. Inoculation of lactic acid bacterial strains in maize silage resulted in differences in nutritional value or population of pathogenic and destructive microorganisms. Strains 16 and 7, identified as Lactobacillus fermentum and Lactobacillus salivarius, were considered promising sources for use as inoculants in maize silages because they provide silages with better fermentation properties and improve aerobic stability after exposure to air.