发病机制的类型和原理Escherichiacoli_部分53

pic gene in strain 042 increased fitness for growth in mucus scraped from the surface of the mouse intestine. Moreover, pic mutants in EAEC are less adept at colonizing the mouse intestine, whereas a Shigella pic mutant elicited less intense intestinal inflammation in the rabbit ligated-loop model (Henderson and Nataro, 2001).
The ShET1 (55-kDa) mode of action has not been defined, but it does not appear to act via the traditional mechanisms of toxin-induced intestinal secre-tion, such as via cyclic AMP and cyclic GMP. ShET1 induces fluid secretion in mucosal tissue explants but does not induce cytotoxic effects (Fasano et al., 1997). Studies have suggested that most EAEC strains from patients with diar-rhea express the ShET1 toxin (Czeczulin et al., 1999; Vila et al., 2000). ShET1 may contribute to the secretory diarrhea that accompanies EAEC and Shigella infections (Kaper et al., 2004).
SepA, a SPATE toxin, was originally described in Shige lla fle xne ri 2a, where it is among the most abundant secreted proteins. The protease is encoded on the Shigella virulence plasmid, suggesting a contribution to pathogenicity (Benjelloun-Touimi et al., 1995). Using human colonic tissue, it was shown that purified SepA toxin elicits mucosal damage (Coron et al., 2009). SepA mutants of S. flexneri induced less mucosal inflammation in ligated rabbit ileal loops compared with the wild-type parent strain (Benjelloun-Touimi et al., 1998). The precise mode of action of SepA remains to be discovered. We have found SepA to be common and epidemiologically important among EAEC strains: among EAEC strains isolated from a case-control study of children’s diarrhea in Mali, SepA was the only factor strongly associated with diarrheal illness (Boisen et al., 2012).
The first EAEC virulence factor that was implicated as a potential cause of diarrhea was the enteroaggregative heat-stable toxin, EAST1. Epidemiological studies demonstrate that EAST1 is not only associated with EAEC but is also present in a wide range of pathogenic E. coli such as ETEC, DAEC, entero-pathogenic E. coli (EPEC), and enterohemorrhagic E. coli. Furthermore, E. coli strains harboring no known virulence factors other than EAST1 were found in the feces of humans with diarrhea (Paiva de Sousa and Dubreuil, 2001; Menard and Dubreuil, 2002). EAST1 is encoded by the astA gene which spans 117 bp. It is a 38-aminoacid peptide with homology to the heat-stable (ST) enterotoxin of ETEC (Savarino et al., 1991, 1993). The astA gene can be found on either plasmids or on the chromosome, and sometimes both, in one or several copies (Menard and Dubreuil, 2002). EAST1 is immunologically different from STa, as no cross-neutralization was observed with polyclonal anti-STa antibodies (Savarino et al., 1996). It is conceivable that EAST1 could contribute to watery diarrhea in EAST1-positive EAEC strains (41% of EAEC strains harbor the astA gene). However, the astA gene is also present in up to 38% of commensal E. coli strains (Savarino et al., 1996; Yamamoto and Echeverria, 1996; Zhou et al., 2002). EAST1 may exist as a series of allelic variants, some of which may be more virulent than others (Menard et al., 2004).
We have described in EAEC several putative pathogenicity islands that may encode additional virulence factors, as predicted from in silico homology to better characterized systems. Some EAEC strains encode a type 3 secretion system called ETT-2; putative effectors for this system were found elsewhere on the genome (Sheikh et al., 2006). Moreover, we have shown that these putative effectors are under transcriptional control of a factor called EilA, homologous to the HilA activator from Salmonella strains. EilA also activates the bacterial surface protein Air, which features predicted immunoglobulin-like repeats. This new putative virulence-related regulon in EAEC may include adherence and aggregation (Sheikh et al., 2006).
INFLAMMATION IN EAEC PATHOGENESIS
EAEC is an inflammatory pathogen, as demonstrated both in clinical (Green-berg et al., 2002) and laboratory (Steiner et al., 1998) reports. Clinical stud-ies have shown that lactoferrin, IL-8, and IL-β can be detected in feces from cases of EAEC diarrhea at a higher level than in stools of patients infected with non-EAEC diarrhea (Jiang et al., 2002). The virulence factors of typical EAEC (including aggA, aggR, aafA, and aap) are associated with increased levels of fecal cytokines and inflammatory markers, and may be observed whether or not the patient manifests diarrhea. The inflammatory effect was linked to expres-sion of a novel EAEC flagellin protein (Donnelly and Steiner, 2002), which is homologous to a flagellin encoded by S. dysenteriae. The EAEC flagellin induced IL-8 from intestinal epithelial cells (IECs) in culture (Steiner et al., 1997, 1998).
It was shown that flagellin was the major pro-inflammatory factor of EAEC on intestinal epithelial cells in culture (Okhuysen and Dupont, 2010). Infec-tion of polarized monolayers of the human colonic intestinal cell line T84 with EAEC strain 042 caused both IL-8 release (Harrington et al., 2005) and a drop in trans-epithelial electric resistance (TEER) when compared with the unin-fected control and with non-pathogenic E. coli HS (Strauman et al., 2010). It is now confirmed in vitro that the fimbriae mediate release of IL-8 and drop in TEER. Furthermore it was shown that AAF/II fimbriae are sufficient to induce transmigration of neutrophils across an epithelial layer in vitro (Qadri et al., 1994). Since a suitable animal model is not in use, a xenotransplant model was used and it was shown that the aggregative fimbriae are important in the devel-opment of inflammation in the human intestine. These data suggest that the AAF adhesins may be not only colonization factors, but may also be both nec-essary and sufficient for induction of mucosal inflammation (Boll et al., 2012). STRAIN HETEROGENEITY
EAEC strains belong to a diverse range and combination of O:H serotypes (Vial et al., 1988; Yamamoto et al., 1992; Qadri et al., 1994; Huppertz et al.,
1997; Olesen et al., 2005; Boisen et al., 2012). Moreover, a high frequency of EAEC strains expresses untypable O antigens and H antigens or are non-motile (Nataro and Kaper, 1998; Uber et al., 2006; Regua-Mangia et al., 2009). Never-theless, there exist commonly isolated EAEC serotypes (O44:H18, O111:H12, O125, and O126 strains). Studies show that some serotypes of the traditional enteropathogenic E. coli such as O55, O111, O86, O126, and O128 can be found in EAEC (Nataro et al., 1998; Suzart et al., 1999; Elias et al., 2002) although the most commonly found serogroups reported in EAEC are O86, O126, and O125 (Spencer et al., 1999; Sarantuya et al., 2004; Uber et al., 2006). It was suggested that the occurrence of EPEC O serogroups (O126, O128, and O158) along with EAEC markers influenced the positive association of E. coli strain with diarrhea (Pereira et al., 2007). Serotyping is often useful in the characterization of other pathogenic E. coli, however it is of little value in EAEC diagnostics (Okeke and Nataro, 2001). However, in a detailed study on genomic characterization by Boisen et al. (2012), targeting a collection of 121 EAEC strains isolated from children in Mali with or without moderate to severe diarrhea, it was found that strains expressing the H33 flagellar antigen were found significantly more often in cases than in controls. This association may signify the existence of a specific set of virulence genes in strains of this H type (Boisen et al., 2012). A phyloge-netic framework was presented identifying three major clusters of DEC contain-ing EAEC. Members of each group show conserved plasmids and chromosomal loci, which indicates that most EAEC, like EPEC, feature conserved linkage of virulence genes (Czeczulin et al., 1999). EAEC strains that did not fall into the above-mentioned clusters could be milder pathogens eliciting inflammation without diarrhea (Steiner et al., 1998). Notably, the pAA plasmid of EAEC is, however, heterogeneous with regard to fimbria and toxin expression. In volun-teer studies, three strains expressing the AAF/I variant did not induce diarrhea, whereas the one strain expressing AAF/II caused diarrhea in the majority of infected subjects (Nataro et al., 1995).
IDENTIFICATION OF EAEC
The gold standard for identifying EAEC is the HEp2-cell adherence assay (Nataro and Kaper, 1998) in as much as the pathogen was initially defined by the presence of a characteristic stacked brick pattern, designated aggregative adherence (AA) in this assay (Nataro et al., 1987). The HEp2-cell adherence assay is unfortunately not designed to screen large numbers of colonies from stool samples as it is very time consuming and is further limited by the risk of contamination of the cell cultures. A DNA probe, CVD432 from the pAA plas-mid of EAEC, has been reported to be specific for EAEC but varies in sensi-tivity (Baudry et al., 1990). As reviewed, the sensitivity variation was between 20% to 89% when compared to the HEp2-cell adherence assay (Okeke, 2009). The CVD432 probe has been shown to correspond to the aatA gene, which encodes a transporter for the dispersin protein (Aap), also regulated by AggR
(Nishi et al., 2003). Aap of EAEC is secreted by many EAEC strains and was sug-gested as a possible target for diagnosis (Sheikh et al., 2002). However, a recent report demonstrates that Aap is also produced by non-EAEC strain. A multiplex PCR assay was developed that detects the three AA plasmid-borne genes (aatA, aggR, and aap) and studies showed that these loci are commonly but not invari-ably linked (Opintan et al., 2010). The authors of this study found that 82% of the EAEC strains isolated from patients with diarrhea were positive for the three loci and that use of a multiplex assay increases both the sensitivity and the specific-ity of EAEC detection (Cerna et al., 2003). Several studies have applied PCR in detecting EAEC targeting genes aggR and/or aatA (Kahali et al., 2004; Sarantuya et al., 2004; Pereira et al., 2007; Regua-Mangia et al., 2009; Gomez-Duarte et al., 2010; Opintan et al., 2010; Rugeles et al., 2010). The variable results obtained using molecular diagnostics may be due to the heterogeneity in EAEC pathogenic mechanisms (Okeke and Nataro, 2001). PCR targeting both virulence factors on the pAA plasmid and the chromosomal EAEC loci, such as aaiC (Dudley et al., 2006) might prove to be the most advantageous approach in detecting EAEC. REFERENCES
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