Bacteriocin diversity, function, discovery and application as antimicrobials.

Bacteriocins are potent antimicrobial peptides that are produced by bacteria. Since their discovery almost a century ago, diverse peptides have been discovered and described, and some are currently used as commercial food preservatives. Many bacteriocins exhibit extensively post-translationally modified structures encoded on complex gene clusters, whereas others have simple linear structures. The molecular structures, mechanisms of action and resistance have been determined for a number of bacteriocins, but most remain incompletely characterized. These gene-encoded peptides are amenable to bioengineering strategies and heterologous expression, enabling metagenomic mining and modification of novel antimicrobials. The ongoing global antimicrobial resistance crisis demands that novel therapeutics be developed to combat infectious pathogens. New compounds that are target-specific and compatible with the resident microbiota would be valuable alternatives to current antimicrobials. As bacteriocins can be broad or narrow spectrum in nature, they are promising tools for this purpose. However, few bacteriocins have gone beyond preclinical trials and none is currently used therapeutically in humans. In this Review, we explore the broad diversity in bacteriocin structure and function, describe identification and optimization methods and discuss the reasons behind the lack of translation beyond the laboratory of these potentially valuable antimicrobials.

Nisin E Is a Novel Nisin Variant Produced by Multiple Streptococcus equinus Strains.

Nisin A, the prototypical lantibiotic, is an antimicrobial peptide currently utilised as a food preservative, with potential for therapeutic applications. Here, we describe nisin E, a novel nisin variant produced by two Streptococcus equinus strains, APC4007 and APC4008, isolated from sheep milk. Shotgun whole genome sequencing and analysis revealed biosynthetic gene clusters similar to nisin U, with a unique rearrangement of the core peptide encoding gene within the cluster. The 3100.8 Da peptide by MALDI-TOF mass spectrometry, is 75% identical to nisin A, with 10 differences, including 2 deletions: Ser29 and Ile30, and 8 substitutions: Ile4Lys, Gly18Thr, Asn20Pro, Met21Ile, His27Gly, Val32Phe, Ser33Gly, and Lys34Asn. Nisin E producing strains inhibited species of Lactobacillus, Bacillus, and Clostridiodes and were immune to nisin U. Sequence alignment identified putative promoter sequences across the nisin producer genera, allowing for the prediction of genes in Streptococcus to be potentially regulated by nisin. S. equinus pangenome BLAST analyses detected 6 nisin E operons across 44 publicly available genomes. An additional 20 genomes contained a subset of nisin E transport/immunity and regulatory genes (nseFEGRK), without adjacent peptide production genes. These genes suggest that nisin E response mechanisms, distinct from the canonical nisin immunity and resistance operons, are widespread across the S. equinus species. The discovery of this new nisin variant and its immunity determinants in S. equinus suggests a central role for nisin in the competitive nature of the species.

Actinomyces Produces Defensin-Like Bacteriocins (Actifensins) with a Highly Degenerate Structure and Broad Antimicrobial Activity.

We identified a strain of Actinomyces ruminicola which produces a potent bacteriocin with activity against a broad range of Gram-positive bacteria, many of which are pathogenic to animals and humans. The bacteriocin was purified and found to have a mass of 4,091 ± 1 Da with a sequence of GFGCNLITSNPYQCSNHCKSVGYRGGYCKLRTVCTCY containing three disulfide bridges. Surprisingly, near relatives of actifensin were found to be a series of related eukaryotic defensins displaying greater than 50% identity to the bacteriocin. A pangenomic screen further revealed that production of actifensin-related bacteriocins is a common trait within the genus, with 47 being encoded in 161 genomes. Furthermore, these bacteriocins displayed a remarkable level of diversity with a mean amino acid identity of only 52% between strains/species. This level of redundancy suggests that this new class of bacteriocins may provide a very broad structural basis on which to deliver and design new broad-spectrum antimicrobials for treatment of animal and human infections.IMPORTANCE Bacteriocins (ribosomally produced antimicrobial peptides) are potential alternatives to current antimicrobials given the global challenge of antimicrobial resistance. We identified a novel bacteriocin from Actinomyces ruminicola with no previously characterized antimicrobial activity. Using publicly available genomic data, we found a highly conserved yet divergent family of previously unidentified homologous peptide sequences within the genus Actinomyces with striking similarity to eukaryotic defensins. These actifensins may provide a potent line of antimicrobial defense/offense, and the machinery to produce them could be used for the design of new antimicrobials given the degeneracy that exists naturally in their structure.

The Lactobacillus casei Group: History and Health Related Applications.

The Lactobacillus casei group (LCG), composed of the closely related Lactobacillus casei, Lactobacillus paracasei, and Lactobacillus rhamnosus are some of the most widely researched and applied probiotic species of lactobacilli. The three species have been extensively studied, classified and reclassified due to their health promoting properties. Differentiation is often difficult by conventional phenotypic and genotypic methods and therefore new methods are being continually developed to distinguish the three closely related species. The group remain of interest as probiotics, and their use is widespread in industry. Much research has focused in recent years on their application for health promotion in treatment or prevention of a number of diseases and disorders. The LCG have the potential to be used prophylactically or therapeutically in diseases associated with a disturbance to the gut microbiota. The group have been extensively researched with regard to stress responses, which are crucial for their survival and therefore application as probiotics.

Recent advances in microbial fermentation for dairy and health.

Microbial fermentation has been used historically for the preservation of foods, the health benefits of which have since come to light. Early dairy fermentations depended on the spontaneous activity of the indigenous microbiota of the milk. Modern fermentations rely on defined starter cultures with desirable characteristics to ensure consistency and commercial viability. The selection of defined starters depends on specific phenotypes that benefit the product by guaranteeing shelf life and ensuring safety, texture, and flavour. Lactic acid bacteria can produce a number of bioactive metabolites during fermentation, such as bacteriocins, biogenic amines, exopolysaccharides, and proteolytically released peptides, among others. Prebiotics are added to food fermentations to improve the performance of probiotics. It has also been found that prebiotics fermented in the gut can have benefits that go beyond helping probiotic growth. Studies are now looking at how the fermentation of prebiotics such as fructo-oligosaccharides can help in the prevention of diseases such as osteoporosis, obesity, and colorectal cancer. The potential to prevent or even treat disease through the fermentation of food is a medically and commercially attractive goal and is showing increasing promise. However, the stringent regulation of probiotics is beginning to detrimentally affect the field and limit their application.