The seed germination process during sprout production provides suitable environmental conditions for the growth of pathogenic bacteria, such as Listeria monocytogenes. A potential way to control this bacterial growth is through the use of bacteriophages, which are naturally occurring viruses that specifically attack bacterial targets and have been shown to be effective antimicrobials in some foods. Therefore, the objective of this study was to evaluate the antimicrobial susceptibility of L. monocytogenes to bacteriophage on alfalfa sprouts during seed germination and subsequent refrigerated storage at 4 °C. Alfalfa sprout seeds were dip-inoculated with 5.5 x 105 CFU/ml L. monocytogenes serogroups 1 and 4. This was followed by treatment with the commercial bacteriophage LISTEX™ P100 at a concentration of 5.3 x 107 PFU/ml. The seeds were then soaked and germinated for 80 h using the glass jar method. The concentration of L. monocytogenes was determined every 24 h using PALCAM agar plated in triplicate. When compared to the spiked, untreated control, treatment of sprout seeds with LISTEX™ P100 resulted in a statistically significant (p < 0.05) reduction of 1.6 log10 CFU/g L. monocytogenes after the initial 24 h of germination. However, the bacteriophage did not show a lasting inhibitory effect, with no statistically significant reductions in L. monocytogenes growth as compared to the control at subsequent time points. The bacteriophage remained stable over the entire germination and storage period. Although biocontrol of Listeria with bacteriophages has high potential to serve as an alternative strategy to control foodborne illnesses, factors such as phage delivery and dose optimization in sprouts need to be further investigated.