Effects of fermented extruded rye flour supplements with Lactobacillus sakei KTU05-6 or Pediococcus pentosaceus BaltBio02 on milk production and composition, as well as ruminal parameters, were determined in Lithuanian Black & White dairy cows. Also, determination of antimicrobial activities of tested lactic acid bacteria (LAB) against a variety of pathogenic and opportunistic bacterial strains previously isolated from diseased cattle was performed. The highest antimicrobial activity was demonstrated in L. sakei against S. aureus, and in P. pentosaceus against P. aeruginosa and S. aureus. The count of LAB in the supplements after 72 h of fermentation of extruded rye flour with L. sakei and P. pentosaceus was 9.6±0.4 log10 CFU/g and 9.5±0.3 log10 CFU/g, respectively. All cows (n=60) were fed the same basal diet. The treatment differences were achieved by individually incorporating (65 d.) one of the supplements: L. sakei KTU05-6 (group B; n=20) or P. pentosaceus BaltBio02 (group C; n=20). The control group A (n=20) was on the basal diet only. A supplement fermented with L. sakei does not have a significant influence on dairy cattle milk production and rumen fluid parameters. The type of LAB used has a significant influence (p<0.0001) on microbiological parameters of the rumen (TCM, TCL, TCE). The milk yield was increased (p≤0.05) using P. pentosaceus BaltBio02 supplement, and further research is needed to identify w hat is the main mechanism of the positive action.