**5. Enteroaggregative** *E. coli* **(EAEC)**

## **5.1 An overview of EAEC**

Enteroaggregative *E. coli* (EAEC) is implicated in epidemic diarrheal illnesses, being a causative agent of traveler's diarrhea, persistent diarrhea in children in EAEC endemic areas, and in immunocompromised patients, particularly in human immunodeficiency virus (HIV) patients [3, 134]. EAEC was first described in 1987 by comparing adherence patterns of *E. coli* isolates to HEp-2 cells, where it showed a stacked-brick aggregative phenotype. EAEC strains are able to infect the colon and/or small bowel of their host where they disrupt the intestinal epithelium and result in loss of electrolytes, watery diarrhea with or without blood and mucus, vomiting, among other symptoms [134]. Persistent EAEC-related diarrhea could result in chronic intestinal inflammation that induces the production of fecal lactoferrin and interleukin (IL-8), and malabsorption of nutrients [135]. Studies on the development of postinfectious irritable bowel disease syndrome in acute EAEC-related diarrhea have been documented [135] but the role of EAEC is not fully understood.

EAEC strains are identified using a molecular probe AA that hybridizes with a region of pAA plasmid encoding an ATP binding cassette transporter apparatus which translocates dispersion across the bacterial cell membrane [136]. Isolates that carry the *aggR* gene that encodes autoagglutination that are associated with persistent diarrhea in patients are able to hybridize with the AA probe. In addition, EAEC has a different adherence pattern, and not all HEp-2 adherent EAEC strains isolated from humans with diarrhea carried *aggR.* Hence, EAEC was classified into two subtypes: typical (*aggR* positive) and atypical (*aggR* negative) subtype [134, 137]. Humans are the reservoir for typical EAEC (tEAEC), whereas atypical EAEC (aEAEC) strains have been isolated from young calves, piglets, and horses as well as companion animals, suggesting the role of animals as a reservoir for this subtype (**Figure 2**) [3].

Although some serotypes including O126:H27, O111:H21, O125, O44:H18 are frequently isolated from EAEC strains, the autoagglutinating phenotype by some EAEC strains complicates the serotyping of this pathotype [3, 138]. In several studies, EAEC strains are often described as nontypeable or as "O?" or O-rough. In a study of EAEC strains from children in Germany, 14 out of 16 isolates that were typeable belonged to different serotypes [139]. Likewise, in a study in the UK, 97 out of 143 EAEC strains that were typeable belonged to more than 40 different O-types [140]. While serotyping is no longer a dependable diagnostic tool for EAEC strains causing diarrheal illness [3, 138], a specific Shiga toxin producing EAEC serotype O104:H4 is associated with a series of outbreaks worldwide [141, 142].

#### **5.2 Virulome of EAEC**

EAEC strains that are implicated in diarrheal illness employ several virulence factors that initiate colonization, promote persistence through adherence to mucosal layers of the intestine, and enterotoxin and cytotoxin secretion (**Table 1**) [134].

#### *5.2.1 Colonization and adherence*

EAEC colonizes the intestinal epithelium of the host using aggregative adhesion fimbriae (AAFs) that also activate the host inflammatory responses and afimbrial adhesins [143]. So far, five AAF variants have been described and are encoded by *aggA* (AAF/I), *aafA* (AAF/II), *agg3A* (AAF/III), *agg4A* (AAF/IV), and *agg5A* (AAF/V) that are regulated by the transcriptional activator *aggR*, borne on EAEC plasmid pAA [143]. AggR also regulates the expression of a type VI Secretion System (T6SS) and a chromosomal PAI encoded by *aaiA*-*aaiP* operon [144] as well all other virulence genes involved in the aggregation and toxin production in pathogenic strains of EAEC (**Figure 3**) [143, 144]. However, in EAEC strains where AAF is absent, an aggregate-forming pili (AFP), a type VI pilus that is encoded by *afp* operon was reported to be responsible for the establishment of a similar aggregative adhesion pattern (**Figure 3**) [144]. Other virulence determinants associated with colonization and adherence of EAEC include *air* gene that codifies for an enteroaggregative immunoglobulin repeat protein and *capU* that encodes a hexosyltransferase homolog, as well as *shf* and *aatA* that have been linked to biofilm formation [3, 144].

#### *5.2.2 Enterotoxin and cytotoxin secretion*

EAEC produces enterotoxins and cytotoxins including EAST1 and colonization factors encoded by *astA* and *pic* genes*,* respectively [37, 145]*.* The latter (*pic*) is often associated with *set1*A and *set1B* encoding two subunits of *Shigella* enterotoxin 1 (ShET1) that are linked to the induction intestinal secretion during infection [145]. This pathotype also carries genes encoding class I cytotoxic SPATE protein family that includes autotransporter proteases encoded by *sigA* and *sepA* and plasmidborne toxin encoded by *pet* [144, 146]. Also, EAEC strains produce dispersin, an anti-aggregation protein that is encoded by *aatPABCD* located on plasmid pAA, and promotes the dispersion of bacteria in the mucosal layer of the intestine [147]. A gene *hlyE* encoding a hemolytic pore-forming toxin that has a cytotoxic effect on cultured cells has also been reported to be present in some EAEC strains, although the role of this gene in the pathogenicity of EAEC is still unclear [3, 148]. Some EAEC strains carry Stx2a phage-encoding Shiga toxin that is associated with HUS in STEC-related infection [141, 146].

The prevalence of the virulence determinants varies with studies and EAEC subtypes [144, 146]. For example, in a study, *pic* gene was reported to be the most prevalent, present in only 47%, while *sepA* and *sigA* were present in less than 15% of the studied isolates [149]. In the study, the authors also noted that *pet* and *pic* genes were associated with tEAEC, whereas *sepA* was associated with aEAEC.

#### **5.3 Antibiotic resistance in EAEC**

EAEC-related diseases such as travelers' diarrhea where antimicrobial therapy is proposed, fluoroquinolones, azithromycin, and rifaximin are often recommended. In immunocompromised patients that require chemoprophylaxis, fluoroquinolones are also considered [150]. For Shiga toxin producing EAEC O104: H4 related infections, azithromycin which has been shown to inhibit *stx* expression in *in-vitro* assay is seldomly used [3, 150].

Although a highly successful treatment rate is achieved with these antibiotics, EAEC strains that are resistant to multiple antibiotics have emerged in different regions [150]. Studies on the resistance of EAEC strains from Southeast Asia,

*The Biology and the Evolutionary Dynamics of Diarrheagenic* Escherichia coli *Pathotypes DOI: http://dx.doi.org/10.5772/intechopen.101567*

India, Africa, and Latin America with travelers' diarrhea showed that more than 50% of the isolates were resistant to ampicillin, sulphamethoxazole, and tetracycline [150, 151]. In a similar study in Iran, 78% and 60% of the extended-spectrum beta-lactamase (ESBL) producing EAEC strains carried the transposable *bla*TEM and *bla*CTX-M genes, respectively [152]. Also, plasmid-mediated quinolone resistance (PMQR) genes (*qnr*) that encode resistance to quinolone have been identified in EAEC in different studies [153, 154]. In England, among the 155 EAEC strains from diarrhea patients in 2015–2016 [155] showing antibiotic-resistant phenotypes, 43 genetic determinants that encode resistance to seven different classes of antibiotics were identified, with *bla*TEM-1 being the most common (40%) followed by *sul2* (37%) and *strA-strB* (32%). Undoubtedly, the rise in antibiotic resistance in this pathotype should be a concern for public health.
