<em>In Silico</em> Identification and Molecular Characterization of <em>Lentilactobacillus hilgardii </em>Antimicrobial Peptides with Activity Against Carbapenem-Resistant <em>Acinetobacter baumannii</em>

This study investigates the efficacy of conditioned media derived from lactic acid bacteria (LAB) in preventing and disrupting Acinetobacter baumannii biofilms. Of the 13 LAB isolates from the Human Microbiome Project tested, five distinct species Lentilactobacillus hilgardii, Lentilactobacillus buchneri, Ligilactobacillus ruminis, Limosilactobacillus fermentum, and Limosilactobacillus antri demonstrated significant prevention of A. baumannii biofilm formation and reduced the biomass of pre-formed mature biofilms. The study extended to the six ESKAPE pathogens and three skin commensals, demonstrating broad-spectrum antimicrobial activity of LAB conditioned media against ESKAPE pathogens, albeit with variable effects on skin commensals. L. hilgardii was characterized further due to its reduced toxicity against skin commensals. Fractionation studies indicated potential peptide involvement; thus, bioinformatic analysis was used to predict antimicrobial peptides (AMPs) within the L. hilgardii genome, which were synthesized and tested, showing effective inhibition of A. baumannii planktonic growth. However, these peptides did not affect biofilms at concentrations up to 30 µg/mL, although synergy with sub-MIC levels of meropenem significantly disrupted mature biofilms. Our study provides a comprehensive platform for the discovery and characterization of AMPs and support the notion of how commensals can be used to reduce, prevent and decolonize biofilms from pathogenic bacteria in community and nosocomial settings.