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The bacterial toxin colibactin causes double-stranded DNA breaks and is associated with the occurrence of bacterially induced colorectal cancer in humans. However, isolation of colibactin is difficult, and its mode of action is poorly understood. Wilson et al. studied Escherichia coli that contain the biosynthetic gene island called pks , which is associated with colibactin production (see the Perspective by Bleich and Arthur). They identified the DNA adducts that resulted from incubating pks + E. coli in human cells. To overcome the lack of colibactin for direct analysis, mimics of the pks product were synthesized. From the resulting synthetic adenine-colibactin adducts, it became evident that alkylation via a cyclopropane “warhead” breaks the DNA strands. Similar DNA adducts were then identified in the gut epithelia of mice infected with pks + E. coli. Science , this issue p. eaar7785 ; see also p. 689 DNA adducts in cells and animals exposed to colibactin-producing gut microbes shed light on the mode of action of a cancer-linked genotoxin. Certain Escherichia coli strains residing in the human gut produce colibactin, a small-molecule genotoxin implicated in colorectal cancer pathogenesis. However, colibactin’s chemical structure and the molecular mechanism underlying its genotoxic effects have remained unknown for more than a decade. Here we combine an untargeted DNA adductomics approach with chemical synthesis to identify and characterize a covalent DNA modification from human cell lines treated with colibactin-producing E. coli . Our data establish that colibactin alkylates DNA with an unusual electrophilic cyclopropane. We show that this metabolite is formed in mice colonized by colibactin-producing E. coli and is likely derived from an initially formed, unstable colibactin-DNA adduct. Our findings reveal a potential biomarker for colibactin exposure and provide mechanistic insights into how a gut microbe may contribute to colorectal carcinogenesis.

Various bacteria are suggested to contribute to colorectal cancer (CRC) development 1 , 2 , 3 , 4 – 5 , including pks + Escherichia coli , which produces the genotoxin colibactin that induces characteristic mutational signatures in host epithelial cells 6 . However, it remains unclear how the highly unstable colibactin molecule is able to access host epithelial cells to cause harm. Here, using the microbiota-dependent ZEB2-transgenic mouse model of invasive CRC 7 , we demonstrate that the oncogenic potential of pks + E. coli critically depends on bacterial adhesion to host epithelial cells, mediated by the type 1 pilus adhesin FimH and the F9 pilus adhesin FmlH. Blocking bacterial adhesion using a pharmacological FimH inhibitor attenuates colibactin-mediated genotoxicity and CRC exacerbation. We also show that allelic switching of FimH strongly influences the genotoxic potential of pks + E. coli and can induce a genotoxic gain-of-function in the probiotic strain Nissle 1917. Adhesin-mediated epithelial binding subsequently allows the production of the genotoxin colibactin in close proximity to host epithelial cells, which promotes DNA damage and drives CRC development. These findings present promising therapeutic routes for the development of anti-adhesive therapies aimed at mitigating colibactin-induced DNA damage and inhibiting the initiation and progression of CRC, particularly in individuals at risk for developing CRC. The oncogenic potential of pks + Escherichia coli depends critically on bacterial adhesion to host epithelial cells mediated by the type 1 pilus adhesin FimH and the F9 pilus adhesin FmlH.

Although Escherichia coli Nissle 1917 (EcN) has been used therapeutically for over a century, the determinants of its probiotic properties remain elusive. EcN produces two siderophore-microcins (Mcc) responsible for an antagonistic activity against other Enterobacteriaceae. EcN also synthesizes the genotoxin colibactin encoded by the pks island. Colibactin is a virulence factor and a putative pro-carcinogenic compound. Therefore, we aimed to decouple the antagonistic activity of EcN from its genotoxic activity. We demonstrated that the pks-encoded ClbP, the peptidase that activates colibactin, is required for the antagonistic activity of EcN. The analysis of a series of ClbP mutants revealed that this activity is linked to the transmembrane helices of ClbP and not the periplasmic peptidase domain, indicating the transmembrane domain is involved in some aspect of Mcc biosynthesis or secretion. A single amino acid substitution in ClbP inactivates the genotoxic activity but maintains the antagonistic activity. In an in vivo salmonellosis model, this point mutant reduced the clinical signs and the fecal shedding of Salmonella similarly to the wild type strain, whereas the clbP deletion mutant could neither protect nor outcompete the pathogen. The ClbP-dependent antibacterial effect was also observed in vitro with other E. coli strains that carry both a truncated form of the Mcc gene cluster and the pks island. In such strains, siderophore-Mcc synthesis also required the glucosyltransferase IroB involved in salmochelin production. This interplay between colibactin, salmochelin, and siderophore-Mcc biosynthetic pathways suggests that these genomic islands were co-selected and played a role in the evolution of E. coli from phylogroup B2. This co-evolution observed in EcN illustrates the fine margin between pathogenicity and probiotic activity, and the need to address both the effectiveness and safety of probiotics. Decoupling the antagonistic from the genotoxic activity by specifically inactivating ClbP peptidase domain opens the way to the safe use of EcN.

Colibactin is a secondary metabolite encoded by the pks gene island identified in several Enterobacteriaceae, including some pathogenic Escherichia coli (E. coli) commonly enriched in mucosal tissue collected from patients with inflammatory bowel disease and colorectal cancer. E. coli harboring this biosynthetic gene cluster cause DNA damage and tumorigenesis in cell lines and pre-clinical models, yet fundamental knowledge regarding colibactin function is lacking. To accurately assess the role of pks+ E. coli in cancer etiology, the biological mechanisms governing production and delivery of colibactin by these bacteria must be elucidated. In this review, we will focus on recent advances in our understanding of colibactin’s structural mode-of-action and mutagenic potential with consideration for how this activity may be regulated by physiologic conditions within the intestine.

Four new dimeric sorbicillinoids (1–3 and 5) and a new monomeric sorbicillinoid (4) as well as six known analogs (6–11) were purified from the fungal strain Hypocrea jecorina H8, which was obtained from mangrove sediment, and showed potent inhibitory activity against the tea pathogenic fungus Pestalotiopsis theae (P. theae). The planar structures of 1–5 were assigned by analyses of their UV, IR, HR-ESI-MS, and NMR spectroscopic data. All the compounds were evaluated for growth inhibition of tea pathogenic fungus P. theae. Compounds 5, 6, 8, 9, and 10 exhibited more potent inhibitory activities compared with the positive control hexaconazole with an ED50 of 24.25 ± 1.57 µg/mL. The ED50 values of compounds 5, 6, 8, 9, and 10 were 9.13 ± 1.25, 2.04 ± 1.24, 18.22 ± 1.29, 1.83 ± 1.37, and 4.68 ± 1.44 µg/mL, respectively. Additionally, the effects of these compounds on zebrafish embryo development were also evaluated. Except for compounds 5 and 8, which imparted toxic effects on zebrafish even at 0.625 μM, the other isolated compounds did not exhibit significant toxicity to zebrafish eggs, embryos, or larvae. Taken together, sorbicillinoid derivatives (6, 9, and 10) from H. jecorina H8 displayed low toxicity and high anti-tea pathogenic fungus potential.

Amphidinols (AM) are a diverse group of bioactive polyketides produced by dinoflagellates of the genus Amphidinium, known for their hemolytic, antifungal, and cytotoxic activities. This work presents the assessment of AM profiles in a comprehensive number of strains, whose species boundaries were previously established through detailed taxonomic analysis. Using UHPLC-MS/MS, we characterized the spectrum of AM analogs in 54 Amphidinium strains isolated from diverse geographical locations. In addition, toxicity was assessed using brine shrimp assays, which revealed significant inter- and intraspecific variability. Despite the broad diversity in AM content, no clear correlation was observed between total AM levels and toxicity across all strains. Multivariate analysis grouped the strains into clusters distinguished by distinct AM profiles and toxicity levels, suggesting that AM production alone does not predict toxicity. Our findings highlight the complexity of Amphidinium bioactivity, emphasizing the influence of strain-specific factors and other bioactive compounds. This work highlights the importance of integrating chemical, genetic, and biological assessments to understand better the factors that govern toxicity in this genus, with implications for ecological studies and the monitoring of harmful dinoflagellates.

An increasing number of human beings from developed countries are colonized by Escherichia coli strains producing colibactin, a genotoxin suspected to be associated with the development of colorectal cancers. Deoxynivalenol (DON) is the most prevalent mycotoxin that contaminates staple food—especially cereal products—in Europe and North America. This study investigates the effect of the food contaminant DON on the genotoxicity of the E. coli strains producing colibactin. In vitro , intestinal epithelial cells were coexposed to DON and E. coli producing colibactin. In vivo , newborn rats colonized at birth with E. coli producing colibactin were fed a DON-contaminated diet. Intestinal DNA damage was estimated by the phosphorylation of histone H2AX. DON exacerbates the genotoxicity of the E. coli producing colibactin in a time- and dose-dependent manner in vitro . Although DON had no effect on the composition of the gut microbiota, and especially on the number of E. coli , a significant increase in DNA damage was observed in intestinal epithelial cells of animals colonized by E. coli strains producing colibactin and coexposed to DON compared to animals colonized with E. coli strains unable to produce colibactin or animals exposed only to DON. In conclusion, our data demonstrate that the genotoxicity of E. coli strains producing colibactin, increasingly present in the microbiota of asymptomatic human beings, is modulated by the presence of DON in the diet. This raises questions about the synergism between food contaminants and gut microbiota with regard to intestinal carcinogenesis. IMPORTANCE An increasing number of human beings from developed countries are colonized by Escherichia coli strains producing colibactin, a genotoxin suspected to be associated with the development of colorectal cancers. Deoxynivalenol (DON) is the most prevalent mycotoxin that contaminates staple food—especially cereal products—in Europe and North America. Our in vitro and in vivo results demonstrate that the intestinal DNA damage induced by colibactin-producing E. coli strains was exacerbated by the presence of DON in the diet. This raises questions about the synergism between food contaminants and gut microbiota with regard to intestinal carcinogenesis. An increasing number of human beings from developed countries are colonized by Escherichia coli strains producing colibactin, a genotoxin suspected to be associated with the development of colorectal cancers. Deoxynivalenol (DON) is the most prevalent mycotoxin that contaminates staple food—especially cereal products—in Europe and North America. Our in vitro and in vivo results demonstrate that the intestinal DNA damage induced by colibactin-producing E. coli strains was exacerbated by the presence of DON in the diet. This raises questions about the synergism between food contaminants and gut microbiota with regard to intestinal carcinogenesis.

Sepsis is a life-threatening infection. Escherichia coli is the first known cause of bacteremia leading to sepsis. Lymphopenia was shown to predict bacteremia better than conventional markers of infection. The pks genomic island, which is harbored by extraintestinal pathogenic E. coli (ExPEC) and encodes the genotoxin colibactin, is epidemiologically associated with bacteremia. To investigate a possible relationship between colibactin and lymphopenia, we examined the effects of transient infection of lymphocytes with bacteria that were and those that were not producing the genotoxin. A mouse model of sepsis was used to compare the virulence of a clinical ExPEC isolate with its isogenic mutant impaired for the production of colibactin. We observed that colibactin induced double-strand breaks in the DNA of infected lymphocytes, leading to cell cycle arrest and to cell death by apoptosis. E. coli producing colibactin induced a more profound lymphopenia in septicemic mice, compared with the isogenic mutant unable to produce colibactin. In a sepsis model in which the mice were treated by rehydration and antibiotics, the production of colibactin by the bacteria was associated with a significantly lower survival rate. In conclusion, we demonstrate that production of colibactin by E. coli exacerbates lymphopenia associated with septicemia and could impair the chances to survive sepsis.

The genotoxin colibactin is a secondary metabolite produced by a variety of pathogenic Enterobacteria, and is associated with colon cancer development and acute systemic infections. The colibactin biosynthesis requires the enzymatic activity of the phosphopantetheinyl transferase ClbA. We recently evidenced that two master regulators of bacterial iron homeostasis, i.e. the ferric uptake regulator (Fur) and the small regulatory non-coding RNA RyhB, were involved in the regulation of the clbA transcription and of the colibactin production. In this study, we investigated the impact of high iron supply on clbA transcription and colibactin production in wild type, Delta ryhB, Delta fur and Delta ryhB Delta fur strains. This revealed that high iron resulted in decreased synthesis of the genotoxin colibactin through both pathways dependent and independent of Fur/RyhB. This work highlights the complex regulatory mechanism that controls an important bacterial virulence and carcinogenesis factor by regulators of bacterial iron homeostasis.

Humans are frequently exposed to bacterial genotoxins involved in digestive cancers, colibactin and Cytolethal Distending Toxin (CDT), the latter being secreted by many pathogenic bacteria. Our aim was to evaluate the effects induced by these genotoxins on nuclear remodeling in the context of cell survival. Helicobacter infected mice, coculture experiments with CDT- and colibactin-secreting bacteria and hepatic, intestinal and gastric cells, and xenograft mouse-derived models were used to assess the nuclear remodeling in vitro and in vivo. Our results showed that CDT and colibactin induced-nuclear remodeling can be associated with the formation of deep cytoplasmic invaginations in the nucleus of giant cells. These structures, observed both in vivo and in vitro, correspond to nucleoplasmic reticulum (NR). The core of the NR was found to concentrate ribosomes, proteins involved in mRNA translation, polyadenylated RNA and the main components of the complex mCRD involved in mRNA turnover. These structures are active sites of mRNA translation, correlated with a high degree of ploidy, and involve MAPK and calcium signaling. Additional data showed that insulation and concentration of these adaptive ribonucleoprotein particles within the nucleus are dynamic, transient and protect the cell until the genotoxic stress is relieved. Bacterial genotoxins-induced NR would be a privileged gateway for selected mRNA to be preferably transported therein for local translation. These findings offer new insights into the context of NR formation, a common feature of many cancers, which not only appears in response to therapies-induced DNA damage but also earlier in response to genotoxic bacteria.