Diffusely-adhering E. coli (DAEC)

Pathogenesis of DAEC

Initial Adhesion and Cell Interaction

Diffuse Adherence Pattern: DAEC is characterized by its ability to adhere uniformly across the entire surface of epithelial cells, a phenomenon known as diffuse adherence (DA). This distinct pattern is observed on cell lines such as HeLa and HEp-2.

Target Cell Preference: The first step of DAEC pathogenesis occurs when the bacteria attach to intestinal epithelial cells. Research using ex vivo methods demonstrates that DAEC bacteria show superior attachment to M cells which are part of Peyer's patch-associated epithelium compared to the microvilli of Peyer's patch-associated epithelium. The bacteria successfully establish themselves in both human undifferentiated crypt cells and differentiated enterocytes.

Initiation of Pathogenesis: DAEC pathogenesis initiates with its adhesion to intestinal epithelial cells.

Virulence Factors and Molecular Mechanisms

Afa/Dr Adhesins and Receptor Binding: Afa/Dr Adhesins and Receptor Binding: The Afa/Dr family of adhesins which includes fimbrial (Dr, F1845) and afimbrial (Afa) types exists on the outer membrane of DAEC bacteria to enable both DA pattern formation on epithelial cells and intestinal epithelial attachment. The adhesins establish connections with particular receptors that exist on host cells.

Figure 2. General genetic organization of afa/dra/daa operons and assembly of Dr and Afa-III adhesins. (Meza-Segura, 2016)

• Human Decay-Accelerating Factor (hDAF): Most members of the Afa/Dr family recognize hDAF (CD55), a 70 kDa glycoprotein on the cell surface. Binding to hDAF induces its clustering around adhering bacteria, a process requiring c-Src kinase activation. hDAF's physiological function is to control complement-cascade amplification.
• Human Carcinoembryonic Antigen-Related Cell Adhesion Molecules (hCEACAMs): Some Afa/Dr family members also bind to hCEACAMs, a group of immunoglobulin-related glycoproteins involved in intercellular adhesion and cell signaling. This binding can induce the recruitment of various hCEACAM receptors and promote cell membrane finger-like extensions.

Induction of Finger-like Projections and Embedding: DAEC attachment to target host cells induces finger-like projections that extend from the surface of infected epithelial cells, effectively embedding the bacteria without complete internalization. This embedding mechanism is believed to provide protection against host-mediated defense mechanisms, antibiotics, and intestinal clearance by peristalsis.

Genetic Organization of Adhesins: The afa/dra/daa operons which produce Afa, Dr and F1845 adhesins consist of five or more genes (A, B, C, D, E). The operon contains five genes which encode a transcriptional regulator and a chaperone and an usher and an invasion protein and an adhesin structural subunit. The D and E subunits exit the cell through the general secretory pathway to reach the periplasm where chaperones in the periplasm help them fold before usher-mediated polymerization.

Secreted Autotransporter Toxin (Sat): The secreted autotransporter toxin (Sat) functions as a major virulence factor. The sat gene appears at higher rates in DAEC strains which cause diarrhea in children compared to DAEC strains from healthy children.

Flagella: Mobile DAEC strains can induce high levels of IL-8 secretion, with purified flagella from Afa/Dr DAEC capable of inducing IL-8 production via TLR5. Since TLR5 is found on the basolateral membrane of polarized epithelial cells, motile DAEC must disrupt tight junctions to allow flagella access to TLR5 and trigger signaling.

Other Virulence Factors: Some DAEC strains carry genes homologous to those of EPEC and EHEC type three-secretion systems, producing EspA, EspB, and EspD homologs, and inducing pedestal formation. Other documented virulence factors include toxins (e.g., astA), siderophores (fyuA, irp2, iuc, iroN), transport systems (shu, modD), and intra-macrophage survival factors (htrA, dsbA).

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Host Cell Responses and Epithelial Damage

Figure 3. DAEC infection. (Meza-Segura, 2016)

• Inflammatory Response: The inflammatory response starts when DAEC infects epithelial cells which release IL-8 to attract neutrophils toward the infection area. The movement of polymorphonuclear leukocytes (PMNL) results in the production of TNF-α and IL-1β which enhances hDAF expression and clustering
• Epithelial Cell Damage: The infection of DAEC leads to two main types of epithelial cell damage which include microvilli loss and adherens junction disruption and actin network breakdown in intestinal cells. The process of microvilli tip release from infected cells depends on the cell differentiation stage.

DAEC Detection Methods

To overcome the limitations of cell adhesion assays, molecular methods that target genes or operon regions of afa and daa are preferred for DAEC identification. These methods ensure more specific detection of DAEC strains. Early DAEC identification relied on DNA probes. PCR Methods: In recent years, PCR methods have become the preferred and more specific approach for DEC identification, including DAEC. Both single and multiplex PCR methods have been developed for this purpose. The first detection methods used diffuse adherence observation but their non-specific nature required scientists to create molecular detection methods. The development of DNA probes brought better results but PCR methods which detect afaC or daaD genes represent the most precise and dependable approach for DAEC identification.