Microbial dysbiosis has been associated with colon cancer. Metabolic and metagenomic analysis studies suggest that CRC‐associated samples have different species compositions compared to control (Table 1). However, inflammation and microbial dysbiosis may not be sufficient alone to promote tumorigenesis. Complex interactions between established risk factors for cancer including genetics, obesity, dietary intake, and alcohol consumption seem to alter the gut microbiota to contribute to colorectal carcinogenesis24, 25 (Fig. 1).

Individual bacterial species and colon cancer risk

Understanding the bacterial species associated with CRC is essential for developing new, specific, and sensitive molecular tools to help in the early detection of colonic diseases.13 Moreover, such as in genetic tumor profiles, finding abundant microorganisms in tumors might become a routine test to guide patient prognosis and management. A list of relevant bacterial species and its role in colorectal carcinogenesis are detailed hereafter.

Fusobacterium nucleatum Genomic methods, including 16 rDNA and shotgun metagenomics analysis, suggest Fusobacterium nucleatum as a potential bacterium contributing to CRC pathogenesis. Kostic and colleagues were the first to show that F. nucleatum promotes myeloid infiltration of intestinal tumors in ApcMin/+ mice and increases the expression of pro‐inflammatory genes such as Ptgs2(COX‐2), Scyb1(IL8), Il6, Tnf (TNFα), and Mmp3.26 Subsequent studies including fluorescent in situ hybridization (FISH) and quantitative polymerase chain reaction further confirmed the rise of F. nucleatum in the colonic mucosa, tissues, and feces of patients with adenomas or adenocarcinomas compared to healthy individuals.12, 27 These results have been consistent throughout different stages of CRCs and across diverse ethnic populations.12, 27 Recent studies also suggest that F. nucleatum levels are valuable for CRC diagnosis.28 However, Kostic et al. revealed that the abundance of F. nucleatum could be persistent in both healthy and CRC hosts suggesting that F. nucleatum detection alone is not robust enough as a biomarker for CRC.26 Investigation of human colonic tissues has revealed complex mechanisms in which F. nucleatum promotes a pro‐tumorigenic microenvironment. Yang et al. incubated CRC cell lines with F. nucleatum in mice and analyzed their microRNA (miRNA) expression patterns.29 Results demonstrated that F. nucleatum upregulates inflammatory factors and microRNA21 through toll‐like receptors.29 F. nucleatum was also seen to affect the downstream targets of miR21, including the oncoprotein RAS P21 Protein Activator 1 (RASA1) and tumor suppressor programmed cell death protein 4 (Pdcd4).29 Experimental evidence suggests that the aberrant expression of RASA1 in CRC leads to the activation of RAS‐mitogen‐activated protein kinase (MAPK) cascade.30 Therefore, F. nucleatum's upregulation of miR21 is strongly correlated with the activation of the MAPK cascade, which could, in turn, contributes to CRC development29 (Fig. 2a,b). Figure 2 Open in figure viewer PowerPoint Fusobacterium nucleatum induces tumor growth and proliferation. Fusobacterium nucleatum activates NFĸB signaling pathway through toll‐like receptors (TLR). This upregulates the transcription of miR‐21, which is then hypothesized to act in two different mechanisms: (a) activation of RAS which phosphorylates transcription factor ERK thereby increasing growth and proliferation and (b) inhibition of Pdcd4 which is responsible for the inhibition of tumor promotion and progression. [Color figure can be viewed at induces tumor growth and proliferation.activates NFĸB signaling pathway through toll‐like receptors (TLR). This upregulates the transcription of miR‐21, which is then hypothesized to act in two different mechanisms: (a) activation of RAS which phosphorylates transcription factor ERK thereby increasing growth and proliferation and (b) inhibition of Pdcd4 which is responsible for the inhibition of tumor promotion and progression. [Color figure can be viewed at wileyonlinelibrary.com Abed and colleagues have identified that polysaccharide D‐galactors‐b (1‐3)‐N‐acetyl‐D‐galactosamine (Gal‐GalNAc), an early biomarker for colon carcinogenesis, was overexpressed in CRC tissues compared to normal colonic tissues.31 The same study has also noted an increased number of F. nucleatum binding to CRC tissues, as opposed to the normal colon. The high levels of F. nucleatum were related to the high expression of Gal‐GalNAc, which binds to fusobacterial lectin Fap2 in CRC tissues.31 The immunological consequences of F. nucleatum translocation to tumor sites remains unclear. Gur et al.32 have demonstrated that Fap2 of the bacteria facilitates immune evasion and thereby promotes tumor growth and progression (Fig. 3b). This is achieved through binding to T‐cell immunoglobulin and ITIM domain expressed on natural killer cells and T cells.32 Further research targeting the pathological effects of the TGIT‐Fap2 is essential as inhibiting this interaction has potential therapeutic implications in CRCs (Fig. 3). Figure 3 Open in figure viewer PowerPoint Fusobacterium nucleatum. (a) Galactose and N‐acetyl‐D‐galactosamine (Gal‐GalNAc) is overexpressed on tumor cells. This increases the binding of Gal‐GalNAc with F. nucleatum protein FAP‐2. (b) the increase of F. nucleatum in the tumor microenvironment promotes its escape from immune cells. The binding of FAP‐2 with TIGIT (T cell immunoglobulin and ITIM domain) inhibits the function of natural killer cells (purple) and T cells (green). [Color figure can be viewed at Tumor immune tolerance and. (a) Galactose and N‐acetyl‐D‐galactosamine (Gal‐GalNAc) is overexpressed on tumor cells. This increases the binding of Gal‐GalNAc withprotein FAP‐2. (b) the increase ofin the tumor microenvironment promotes its escape from immune cells. The binding of FAP‐2 with TIGIT (T cell immunoglobulin and ITIM domain) inhibits the function of natural killer cells (purple) and T cells (green). [Color figure can be viewed at wileyonlinelibrary.com Experiments by Rubinstein et al.33 demonstrated a relationship between FadA component of F. nucleatum and E‐cadherin, a cell adhesion molecule and a tumor suppressor that functions through the β‐catenin signaling pathway (Fig. 3a). In the presence of F. nucleatum, the FadA portion interacts with E‐cadherin inhibiting its tumor suppression activity and activating β‐catenin signaling pathway in a similar mechanism as that of other growth factors such as endothelial growth factor and vascular endothelial growth factor. This will, in turn, result in the activation of oncogenes, Wnt genes, and inflammatory genes33 (Fig. 4). Figure 4 Open in figure viewer PowerPoint Fusobacterium nucleatum mediates the expression of oncogenes. The interaction between the FadA component of F. nucleatum and E‐cadherin stimulates a carcinogenic pathway that is often induced via endothelial growth factors (EGF) and vascular endothelial growth factors (VEGF). This pathway induces the dissociation of β‐catenin from α‐catenin and translocates it into the nucleus for β‐catenin‐regulated transcription. This results in the activation of oncogenes, Wnt genes, and inflammatory genes, a phenotype that is consistent with adenomatous polyposis coli (APC) mutation. [Color figure can be viewed at mediates the expression of oncogenes. The interaction between the FadA component ofand E‐cadherin stimulates a carcinogenic pathway that is often induced via endothelial growth factors (EGF) and vascular endothelial growth factors (VEGF). This pathway induces the dissociation of β‐catenin from α‐catenin and translocates it into the nucleus for β‐catenin‐regulated transcription. This results in the activation of oncogenes, Wnt genes, and inflammatory genes, a phenotype that is consistent with adenomatous polyposis coli (APC) mutation. [Color figure can be viewed at wileyonlinelibrary.com Recent studies support the potential role of F. nucleatum in mediating the association between diet and colorectal neoplasms.34 Interestingly, foods rich in red and processed meat were associated with F. nucleatum positive tumors but not with F. nucleatum negative tumors.34 Also, this association was stronger for proximal colon cancers compared to distal CRCs.34 The altered levels of F. nucleatum and its association with CRCs across different populations could be attributed to differences in the diet.27 Phenotypically, F. nucleatum has the ability to ferment undigested proteins to produce hydrogen sulfide,35 which could induce carcinogenesis in the colon.36

Streptococcus gallolyticus subsp. gallolyticus (Sgg) Since 1951, clinical studies linked the presence of Streptococcus bovis biotype I, renamed S. gallolyticus subsp. gallolyticus (Sgg), to colon carcinoma.37 Recently, Kumar et al.38 established the tumor‐promoting role of Sgg that involves specific bacterial and host factors. The strong association between Sgg and CRC indicates that the bacterium may have specific pathogenic traits that aid in the promotion and spread of cancer. S. bovis/gallolyticus can adhere to colon epithelial surfaces and can bind to heparin sulfate proteoglycans on malignant cells through its histone‐like protein A39 or to components of the extracellular matrix such as collagen.40 The bacterium pili demonstrate phase‐variable expression, which is an advantageous feature for its colonization around tissues and evasion of the host immune responses.41 The pili are also highly immunogenic and hence anti‐pilins IgG could potentially be an ideal serological diagnostic tool in patients with early adenomas.42 Upon binding to tissues with a malignant phenotype, multiple in vitro experiments demonstrated that S. bovis/gallolyticus proteins stimulate the production of inflammatory cytokines.43 These inflammatory cytokines, in turn, mediate the production of free radicals and thereby the expression of nitric oxide enzyme (NOS2) producing nitric oxide (NO).44, 45 In the colonic epithelial cells, NO was seen to directly regulate oncogenes or tumor suppressor genes to promote its carcinogenic properties.46 The NO, produced by S. bovis/gallolyticus, can activate NFκB, an inflammation‐induced carcinogenesis transcription factor. This mediates angiogenesis through activation of vascular endothelial growth factors, and cellular proliferation through COX‐2 production and c‐Myc and cyclin‐D activation46 (Fig. 5). Figure 5 Open in figure viewer PowerPoint Streptococcus bovis/gallolyticus adheres to the epithelium, initiates inflammation, and promotes angiogenesis and proliferation. The role of Streptococcus bovis/gallolytics in promoting different stages of carcinogenesis is demonstrated. (a) Streptococcus bovis/gallolyticus, through a histone‐like protein A and collagen‐binding proteins, can bind respectively to heparin sulfate proteoglycans on malignant cells and collagens I, II, III, and IV in the extracellular matrix. (b) The adherence of the bacterium to malignant cells stimulates the production of inflammatory cytokines, which induce carcinogenic free radical damage and the expression of inducible nitric oxide enzyme (NOS2) coded by NOS2 genes. (c) The activation of NOS enzymes promotes nitric oxide (NO) production thereby inducing oncogene activation and tumor suppressor gene inactivation. NO also facilitates the production of vascular endothelial growth factors through the activation of NF‐B. (d) NO, bacterial proteins, and inflammatory mediators induce COX‐2 mediated prostaglandins production, which mediates cellular proliferation. The bacterium also increases β‐catenin levels which induce cellular proliferation through c‐Myc and cyclin‐D activation. [Color figure can be viewed at adheres to the epithelium, initiates inflammation, and promotes angiogenesis and proliferation. The role ofin promoting different stages of carcinogenesis is demonstrated. (a), through a histone‐like protein A and collagen‐binding proteins, can bind respectively to heparin sulfate proteoglycans on malignant cells and collagens I, II, III, and IV in the extracellular matrix. (b) The adherence of the bacterium to malignant cells stimulates the production of inflammatory cytokines, which induce carcinogenic free radical damage and the expression of inducible nitric oxide enzyme (NOS2) coded by NOS2 genes. (c) The activation of NOS enzymes promotes nitric oxide (NO) production thereby inducing oncogene activation and tumor suppressor gene inactivation. NO also facilitates the production of vascular endothelial growth factors through the activation of NF‐B. (d) NO, bacterial proteins, and inflammatory mediators induce COX‐2 mediated prostaglandins production, which mediates cellular proliferation. The bacterium also increases‐catenin levels which induce cellular proliferation through c‐Myc and cyclin‐D activation. [Color figure can be viewed at wileyonlinelibrary.com Studies have also revealed that the production of cell proliferation markers following the bacteria colonization increases the risk of colonic adenomas by 50%, which provides further evidence on the role of Sgg “as promoter/propagator of colorectal carcinoma rather than just a consequence of the colorectal tumours.”43 Aymeric et al.47 have suggested that the bacterium benefits from tumor metabolites and can kill closely related enterococci commensals through an SGG‐specific bacteriocin, gallocin, thereby allowing a better colonization niche. Gallocin activity is improved in the presence of secondary bile acids, an established risk factor for CRC.47 It was also shown that the presence of premalignant conditions and APC gene mutation enhances Sgg colonization in a gallocin‐dependent manner.47 In addition, the Wnt pathway activation, one of the earliest signaling alternation in CRC, decreases the expression of bile acid apical transporter gene thereby establishing a new link between Wnt pathway activation and high levels of secondary bile acid.47 Furthermore, Sgg colonization, accompanied by APC mutation and increased carcinogenic secondary bile acids, could potentially be part of a tumorigenesis triangle.42 Recent studies have indicated that Sgg is a passenger and cancer‐promoting bacterium and may thus act as a marker for CRC screening.42 A most recent study by Thomas et al.13 have further confirmed S. gallolyticus as a potential marker for CRC. However, because it requires premalignant conditions for colonization, Sgg might not induce carcinogenesis but rather accelerates its pathogenic progression.42 Also, the evidence of the bacteria in the transformation of aberrant crypts to adenoma and cancer could aid in the early detection of colorectal lesions and thereby act as a potential target in colon carcinogenesis.42, 47

Escherichia coli Certain Escherichia coli strains including B2 genotoxic E. coli, and enteropathogenic E. coli, or tightly adherent E. coli frequently colonize the colorectal mucosa.48 During carcinogenesis, the colonic epithelial changes mainly the loss of tight junctions increase the internalization of E. coli to the tumor tissues.49 Inflammatory mediators, including interferon and tumor necrosis factor, in the carcinogenic microenvironment, upregulate the expression of carcinoembryonic antigen‐related cell adhesion molecule 6.49 Upregulation of carcinoembryonic antigen‐related cell adhesion molecule 6 could lead to the adhesion, invasion, and multiplication of E. coli type 1 pili in the colonic epithelium,49 which further promotes tumor survival. Arthur et al.14 provided the first evidence of the prevalence of E. coli strains of B2 phylotype in CRCs compared to healthy individuals. In the B2 phylogroup, the expression of polyketide synthase (pks) island containing genotoxin colibactin peptide is thought to induce double‐strand breaks in DNA.48 The defined genotoxic factors such as cytolethal distending toxin or undefined genotoxic factors49 can also modulate the enterocyte transformation and induce inflammatory reactions in the intestinal epithelium.50 An alternative hypothesis suggests that rather than a carcinogen, colibactin is a bacteriocin that kills commensals, which gives the phylotype a selective advantage.51