Microcins are a class of antimicrobial peptides produced by certain Gram-negative bacterial species to kill or inhibit the growth of competing bacteria. Only 10 unique, experimentally validated class II microcins have been identified, and the majority of these come from Escherichia coli. Although the current representation of microcins is sparse, they exhibit a diverse array of molecular functionalities, uptake mechanisms, and target specificities. This broad diversity from such a small representation suggests that microcins may have untapped potential for bioprospecting peptide antibiotics from genomic data sets. We used a systematic bioinformatics approach to search for verified and novel class II microcins in E. coli and other species within its family, Enterobacteriaceae. Nearly one-quarter of the E. coli genome assemblies contained one or more microcins, where the prevalence of hits to specific microcins varied by isolate phylogroup. E. coli isolates from human extraintestinal and poultry meat sources were enriched for microcins, while those from freshwater were depleted. Putative microcins were found in various abundances across all five distinct phylogenetic lineages of Enterobacteriaceae, with a particularly high prevalence in the "Klebsiella" clade. Representative genome assemblies from species across the Enterobacterales order, as well as a few outgroup species, also contained putative microcin sequences. This study suggests that microcins have a complicated evolutionary history, spanning far beyond our limited knowledge of the currently validated microcins. Efforts to functionally characterize these newly identified microcins have great potential to open a new field of peptide antibiotics and microbiome modulators and elucidate the ways in which bacteria compete with each other. IMPORTANCE Class II microcins are small bacteriocins produced by strains of Gram-negative bacteria in the Enterobacteriaceae. They are generally understood to play a role in interbacterial competition, although direct evidence of this is limited, and they could prove informative in developing new peptide antibiotics. However, few examples of verified class II microcins exist, and novel microcins are difficult to identify due to their sequence diversity, making it complicated to study them as a group. Here, we overcome this limitation by developing a bioinformatics pipeline to detect microcins in silico. Using this pipeline, we demonstrate that both verified and novel class II microcins are widespread within and outside the Enterobacteriaceae, which has not been systematically shown previously. The observed prevalence of class II microcins suggests that they are ecologically important, and the elucidation of novel microcins provides a resource that can be used to expand our knowledge of the structure and function of microcins as antibacterials.
Keywords: Enterobacteriaceae; Gram-negative bacteria; bacteriocins; bioinformatics; computational biology; microcin.