There is an increasing demand for high-quality silage production with improved nutrients as a feed resource for livestock. The addition of lactic acid bacteria (LAB) is one way to enhance silage quality via increasing lactic acid content. The objective of the present study was to isolate and identify potent LAB and investigate their effects on in vitro fermentation in low moisture Italian ryegrass (IRG) silage. LAB strains were isolated from alfalfa (Medicago sativa) using a selective media and then the antifungal activity were assessed by pour plate method; selected LAB were characterized and identified by biochemical and 16S rRNA sequencing. For silage production, low moisture (<50%) IRG was chopped and packed in sterile air-tight polypropylene bags with selected LAB colonies and sealed in an anaerobic condition using a vacuum-sealing machine. After a month, the number of LAB colonies and fermentative metabolites was analyzed in the samples. Statistical analysis was performed on all variables using SPSS/PC. One-way ANOVA and Duncan multiple range tests were used with 5% probability level. Thirty LAB strains were isolated and screened for their antifungal activity against Aspergillus species; among these strains, 2 strains exhibited strong antifungal activity and named as KCC-37 and KCC-38. 16srRNA sequence revealed that KCC-37 and KCC-38 were Lactobacillus plantarum. Both strains were able to survive under harsh conditions of GIT tract. Low moisture IRG with either KCC-37 or KCC-38 reduced pH of the silage and improved the organic acids productions than the non-inoculated samples. Overall, these isolated strains could be used as an additive for the improvement of the fermentation quality of the low moisture silage.