The Hungate1000 prokaryotic culture collection encodes a wide variety of bacteriocins

Bacteriocins are antimicrobial peptides/proteins that are widely distributed among bacteria and are gathering traction as natural alternatives to antibiotics, modulators of the microbiota, and interbacterial signaling peptides. The Hungate1000 is a culture collection of isolated prokaryotic microorganisms and their genomes from ruminant animals that aims to expand the knowledge base of rumen ecology. In this study, 410 rumen-isolated prokaryotes within the collection were mined to expand upon the bacteriocin-producing potential of the rumen. A total of 408 novel bacteriocin gene clusters were identified across 308 genomes. Bacteriocins in novel species within the Hungate1000 were identified, such as Pseudobutyrivibrio sp. UC1225, which has two novel natural nisin variants, Clostridium sp. DSM 8431 with a novel peptide 81% identity to amylocyclicin and Lachnobacterium C7 encoding a novel circular bacteriocin with 55% identity to the circular bacteriocin NKR-5-3B. A novel class II lanthipeptide gene cluster was also identified containing eight distinct core peptides encoded within the genome of a novel Butyrivibrio species. Bacteriocin biosynthetic potential was noted within species unknown to produce bacteriocins, such as Lachnobacterium bovis DSM 14045, Lachnospira multipara D15d, Eubacterium callanderi NLAE-zl-G225, Eisenbergiella tayi NLAE-zl-G231, and Muricomes contorta NLAE-zl-C134. The frequency of putative bacteriocin production within ruminal strains was 30%, doubling the frequency previously suggested in the mammalian gastrointestinal tract. This number increases to ~70% when encompassing groups of peptides with limited knowledge of antibacterial activity, such as ranthipeptides and auto-inducing peptides. We also show that the bacteriocin core peptides mined from the Hungate1000 culture collection are found in the microbiomes of other ruminant animals and the human gut microbiome. These findings highlight the Hungate1000 as a rich biosynthetic reservoir of cultured strains that can be experimentally explored for functional antimicrobial activity. The presence of diverse bacteriocin-producing lineages in rumen-associated microbes provides a foundation for future strategies aimed at targeted microbiome modulation, including approaches to improve rumen function and potentially mitigate enteric methane emissions using bacterial strains or their natural products.IMPORTANCEBacteriocins are gathering traction as a possible alternative to antibiotics in some instances. Therefore, it is crucial to discover novel bacteriocins to expand the bacteriocin knowledge base if these peptides are to be translated to the clinic for use in humans or developed as veterinary interventions to modulate rumen function. Here, we use in silico methods to identify the biosynthetic potential of the Hungate1000 culture collection of rumen bacterial strains. We discover 408 novel bacteriocin gene clusters across 308 genomes and identify that the frequency of bacteriocin gene clusters is over 30%, which is double the incidence rate from previous studies of the mammalian gastrointestinal tract. This number increases to approximately 70% when including bacteriocin classes such as ranthipeptides and cyclic-lactone-autoinducer peptides. Together, these findings position the rumen microbiome as a rich and underexplored reservoir of antimicrobial diversity, with potential for the development of targeted microbiome-modulating therapeutics, livestock interventions aimed at improving rumen function, and strategies aligned with One Health goals, including antimicrobial stewardship and methane mitigation.