Bacillus pumilus APC 4184 produces a plethora of antimicrobial metabolites: Insights into pumilacidins and the novel circular bacteriocin pumilarin X

Bacillus species are important in food biopreservation due to their bioactive metabolites – bacteriocins and lipopeptides – which hold great potential to combat foodborne pathogens like Listeria monocytogenes . These metabolites may replace natural alternatives. This study focused on Bacillus pumilus APC 4184, an apricot stick isolate, to identify and characterize its key antimicrobial metabolites. APC 4184 was screened against a range of foodborne pathogens and spoilage bacteria. The anti- Listeria metabolite was further analyzed using the kinetic production study, genome mining, MALDI-TOF mass spectrometry, and heterologous expression in Lactococcus lactis . Pumilacidins, a group of lipopeptides, were characterized by genome mining, partial purification, and antibacterial assessment. Results showed that APC 4184 exhibits a broad inhibitory spectrum, mainly against gram-positive bacteria. The novel circular bacteriocin pumilarin X (7045 Da) was produced during early growth and identified as the anti- Listeria metabolite. Genome analysis revealed the biosynthetic gene clusters of pumilarin X and pumilacidins (molecular masses of 1058, 1072, 1086, and 1100 Da). Purified pumilacidins exhibited specific activity against L. lactis HP but were inactive against Listeria and Staphylococcus . Our study offers valuable insights into the target specificity of Bacillus pumilus metabolites. We identify the traits of pumilarin X and pumilacidins as narrow-spectrum agents. These discoveries allow for the selective extraction of key metabolites from complex mixtures, supporting precise applications of bioactive agents in food systems. • Bacillus pumilus APC 4184, isolated from dry apricot, exhibits broad inhibition spectrum against various gram-positive foodborne pathogens. • Bacillus pumilus APC 4184 produces pumilarin X, a novel circular bacteriocin, early in growth, exhibiting potent anti- Listeria activity. • Pumilarin X was successfully cloned and expressed in Lactococcus lactis (pNZ44-pumilarin X), showing potent antagonistic activity against Listeria and Bacillus species. • Bacillus pumilus APC 4184 also produces pumilacidins, which inhibited the growth of Lactococcus lactis HP.