Bacillus species have recently drawn attention due to their potential use in the biological control of fungal diseases. This paper reports on the antifungal activity of novel peptides isolated from Bacillus amyloliquefaciens PT14. Reverse-phase high-performance liquid chromatography revealed that B. amyloliquefaciens PT14 produces five peptides (PT14-1, -2, -3, -4a, and -4b) that exhibit antifungal activity but are inactive against bacterial strains. In particular, PT14-3 and PT14-4a showed broad-spectrum antifungal activity against Fusarium solani and Fusarium oxysporum. The PT14-4a N-terminal amino acid sequence was identified through Edman degradation, and a BLAST homology analysis showed it not to be identical to any other protein or peptide. PT14-4a displayed strong fungicidal activity with minimal inhibitory concentrations of 3.12 mg/L (F. solani) and 6.25 mg/L (F. oxysporum), inducing severe morphological deformation in the conidia and hyphae. On the other hand, PT14-4a had no detectable hemolytic activity. This suggests PT14-4a has the potential to serve as an antifungal agent in clinical therapeutic and crop-protection applications.