New Antibiotics Discovered from Human Microbiome

The human microbiome refers to the microbial community that lives inside and outside the human body, and although the total weight of the human microbiome is only 500 grams, the ratio of microbial cells to human cells is about 1:1, and the gene content of the microbiome is 150 times the gene content of the human chromosome. As an important "invisible organ", the human microbiome plays a key role in digestion, nutrient synthesis, immune regulation, protection of the gastrointestinal tract and vagina from pathogens, drug metabolism, and regulation of mental health. Bioactive compounds found in human microbes, which inhibit the colonization of competitors (including pathogenic bacteria), play an important role in protecting the body from pathogens and boosting host immunity.

For example, the non-ribosomal cyclic peptide Lugdunin from the nasal cavity of Staphylococcus lugdunensis can effectively inhibit methicillin-resistant Staphylococcus aureus (1.5 μg/mL) without developing resistance. It can also promote the expression and release of LL-37 and CXCL8 and recruit phagocytic cells to enhance the defense capability of human host. Epifadin, produced by bacteria of the genus Staphylococcus in the nasal cavity of healthy humans, has a molecular structure that incorporates peptide, polyene and tetramate groups. Epifadin is a broad spectrum antibiotic, but it is very unstable and has inspired novel antibiotic development strategies because of its potential to reduce side effects on probiotics and normal cells. The probiotic Streptococcus salivus K12, isolated from the mouths of healthy infants, has a 190 kb giant plasmid, pSsK12, which contains two biosynthetic gene clusters (BGCs) encoding two different bacteriotin families. Some strains of this species produce the antibiotic Salvaricins and their phosphorylated products, which are able to penetrate oral cells, kill specific disease-associated bacteria, including Streptococcus pyogenes, and boost host immunity by activating neutrophils and promoting phagocytosis. There is also A trans-AT PSSKS-NRPS BGC (sar) on the plasmid pSsK12 whose salAB (responsible for the biosynthesis of Salvaricins) deletion mutant produces Salivabactin (hybrid 4/5), which is active against Gram-positive pathogens such as Group A Streptococcus (GAS), The protective effect in vivo is close to that of penicillin G.

The above case highlights that human colonizing bacteria can form chemical barriers through peptides and peptide-polyketone hybrid compounds to block pathogen invasion. Through the big data resources provided by the human microbiome project (HMP), studying the biosynthesis of healthy human colonized bacteria can better understand the relationship of the human microbiome in health and disease, and facilitate the development of treatment options. After a systematic review of 752 metagenomes of phase I HMP, 3118 BGCs were identified from healthy individuals, including sugars, non-ribosomal peptides (NRPs), polyketoids (PKs), terpenoids, ribosome modified peptides (RiPPs), and heterozygotes. These BGC-derived bacteria genera are uncommon in non-human environments, such as Bacteroides, Parabacteroides, Corynebacterium, Rothia, and Ruminococcus, highlights the unique biosynthetic capabilities of the human microbiome. This article gives a comprehensive list of relevant cases.

A natural antimicrobial from human nose

Strains of S. epidermidis IVK83 isolated from the nasal cavity of healthy volunteers showed strong inhibition against a variety of bacteria, including methicillin-resistant Staphylococcus aureus. By screening a library of transposon mutants constructed from IVK83 strains, the researchers identified a new BGC consisting of ten genes encoding a variety of biosynthases. When the gene efiA was inactivated, loss of antibacterial activity was observed.

Further studies showed that the culture filtrate of IVK83 strain contained antibacterial active ingredients, which were enriched in DMSO through specific treatment steps and named Epifadin. Epifadin is a new antibacterial substance produced by Staphylococcus epidermidis. Its structure consists of peptide, polyene and tetramine unit. Epifadin showed broad-spectrum inhibition of human nasal microbiota, especially against a variety of gram-positive bacteria and certain actinomycetes. Staphylococcus aureus is particularly sensitive to this compound, while some epidermal staphylococcus strains have shown resistance. In addition, Epifadin also showed inhibitory activity against some fungi. Its mechanism of action mainly involves destroying the integrity of bacterial cell membranes. Analysis of the Staphylococcus aureus genome revealed that specific gene mutations may be associated with its resistance to Epifadin.

Chemical structure of Epifadin. ( Torres Salazar, B. O., 2024)

Antibiotics at BOC Sciences

Antibiotic production services at BOC Sciences

Antimicrobial agents from human vaginal microbiome

Lactobacillus gasseri, one of the four bacteria commonly found in the vaginal microbiome, maintains an acidic vaginal environment by producing lactic acid and synthesizes antimicrobial ribosom-like peptides (RiPPs) to defend against pathogen infections. Lactobacillus JV-V03 has a plasmid encoding thiopeptide BGC and is expressed in wild-type strains to produce Lactocillin. Lactocillin specifically inhibits gram-positive bacteria and has a relatively narrow antibacterial spectrum. Its structural features include indole-S-cysteine, trithiazolylpyridine and multiple thiazole rings, which are formed by post-translational modifications without the involvement of oxygen, and are consistent with the growth conditions of lactic acid bacteria in the anaerobic environment of the vagina. Lactobacillus LEAF 2052A-d, isolated from the vagina of a bacterial vaginosis patient, encodes a BGC(tybA-E). A nucleotide Tyrocitabine with a positive ester link in its phosphoric acid portion was identified by heterogeneously expressing the BGC in Pseudomonas putidis by inserting an orthogonal T7 promoter. Tryocitabine was able to inhibit the translation of the GFP reporter protein, whereas acylated Tyrocitabine lost this biological activity, suggesting a precursor drug mechanism.

Other antimicrobial agents from human microbiome

A computational study of 2229 genomic RiPPs from phase 1 and 2 HMP focused on lanthipeptides and lasso peptides that have specific effects on Gram-positive and Gram-negative bacteria. Twenty-three RiPPs were identified through targeted genome mining and allogeneic expression in E. coli. LANII-687 from Lactobacillus vaginalis LEAF 2053A-b has broad-spectrum antimicrobial activity, particularly against vancomycin-resistant Enterococcus faecalis (VRE) and Staphylococcus aureus. LANII-691 from cutaneous associated Streptococcus pyogenes GA19702 had a similar antibiotic profile but was less potent; Lanthiopeptides from oral Streptococcus sp.M344, such as LANII-286, LANII-287, LANII-916, and LAS-1009 from Roche Bacillus F0474, such as LAS-1009, specifically inhibit the gastrointestinal symbiotic Bifidobacterium adolescentis. Oral bacteria can be transferred to the intestine through saliva, ingestion of contaminated food or blood infection after periodontal disease. They colonize in the intestinal environment and interact with the resident microbial community. This may lead to the decrease of the beneficial symbiotic bacteria Bifidobacterium adolescentis, which may increase intestinal permeability. It forms a pro-inflammatory state and may lead to inflammatory bowel disease. In addition, the altered microbial environment will promote the growth of pathogenic bacteria and carcinogenic processes, promoting the development of colorectal cancer and cirrhosis. Lanthiopeptides and noose peptides targeting the oral bacteria Bifidobacterium adolescentis suggest that RiPPs are involved in pathogenic processes, including intestinal flora dysregulation, primary sclerosing cholangitis and ulcerative colitis. This conclusion partly explains the molecular mechanism of ecological dysbiosis caused by microbial translocation.

The hair follicles and glands of human skin are rich in lipids and proteins, creating an acidic, high-salt, dry, and aerobic niche for symbiotic bacteria to grow, while also potentially harboring opportunistic pathogens such as Staphylococcus aureus. The BGC of the Ripps-like molecule Cutimycin is present in some isolated isolates of Keratobacterium acnes, one of the most common species in the human skin microbiome. Cutimycin is structurally similar to Lactocillin, which inhibits both Staphylococcus aureus and Staphylococcus epidermidis, partially explaining that the overgrowth of S. acneis may be due to Cutimycin's inhibition of S. epidermidis leading to the development of acne, while S. epidermidis controls the proliferation of S. acneis in healthy skin. Cutimycin BGC was also present in nasal microbiome metagenomic data, suggesting that it plays a protective role in niche competition by inhibiting pathogen colonization across body parts.

The human gut microbiota mainly contains a family of gene clusters coding for NRPSs with terminal reductase domains that are responsible for catalytic reduction offloading to produce C-terminal aldehydes. The structure of the product is similar to that of the anticancer drug bortezomib (protease inhibitor), some dipeptide-aldehyde analogues such as Ph-PH-H show strong inhibitory activity against lysosomal cysteine proteinase, while N-acyl Met-PH-H shows nanomolar activity against cathepsin S with unique selectivity.

MetaBGC used in human microbiome studies

In order to directly mine BGC in metagenomic sequencing data of the human microbiome, the MetaBGC algorithm was developed. Using 2544 metagenomic samples from Phase 1 HMP and MetaHIT (Human Intestinal Metagenomics of Human Intestinal), the study aims to identify type II PKS BGCs that are uncommon in the human microbiota. Thirteen complete BGCs were identified, six of which were actively transcribed in humans. Two BGCs from oral actinomyces Rothia dentocariosa and intestinal Blautia wexlerae were hetero-expressed in Streptomyces albus J1074 and Bacillus subtilis, and five aromatic polyketones such as Metamyins and Wexrubicin were identified. Wexrubicin was structurally similar to the anti-cancer drug adriamycin and was not toxic to the human HeLa cell line at tested doses. Metamycin D has strong antibacterial activity against gram-positive bacteria, especially against oral pathogenic bacteria, such as Streptococcus salivarius, Streptococcus intermedia, Streptococcus salivarius and Atopo variolis, suggesting that they have potential roles in oral niche competition and pathogen defense.

References

1. Wang, Y., et al. Exploring nature's battlefield: organismic interactions in the discovery of bioactive natural products. Natural Product Reports. 2024.
2. Torres Salazar, B. O., et al. Commensal production of a broad-spectrum and short-lived antimicrobial peptide polyene eliminates nasal Staphylococcus aureus. Nature Microbiology. 2024, 9(1): 200-213.