A Preliminary Investigation on the Chemical Composition of the Cell Surface of Five Enteropathogenic Escherichia coli Serotypes

Abstract

Enteropathogenic Escherichia coli (EPEC) is a leading cause of diarrhea in children living in developing countries, especially among infants from 6 months to 1 year old (Gomes et al. 1989, Regua et al. 1990, Albert et al. 1995, Tamura et al. 1996. Although diarrhea is often severe and protracted, EPEC virulence factors are not completely well understood (Baldwin 1998).Bacterial adhesion to intestinal epithelial cells has been considered an important step for the pathogenicity of EPEC serotypes leading to ultrastructural and functional changes (Donnenberg et al. 1998). It has been observed that EPEC strains present multifactorial virulence mechanisms and chemical and physical properties of their surfaces are assumed to be related to pathogenicity (Orskov et al. 1977). In nature, bacteria adhere to different substrata by a mass of tangled polysaccharides fibers, a component outside the cell wall (Costerton et al. 1978). This exopolysaccharide network mediates adhesion of bacteria to various substrata, including certain epithelial surfaces of the human host (Peterson & Quie 1981). On the other hand, extracellular capsular polysaccharides of different bacteria, including E. coli, have been frequently associated with diseases (Muhldorfer & Hacker 1994).The aim of the present study was to investigate and compare the chemical nature of exopolysaccharides isolated from different E. coli serotypes. The cellular topology of these components was also assayed by transmission electron microscopy. MATERIALS AND METHODSStrains -EPEC strains belonging to five different serotypes (O111:H2; O111:H12; O125:H9; O119:H6 and O26:H11) were analyzed. They were selected on the basis of the frequency of serotypes isolated during a study previously performed in our laboratory (Regua Mangia et al. 1993). For all experiments, bacteria cells were grown in brain heart infusion broth (BHI; Difco Laboratories, Detroit, Michigan, USA) for 18 hr.Lectin agglutination -Agglutination tests were performed in 96-well, round bottom microtiter plates (Takatsy microtiter plates; Cooke Engineering Co., Alexandria, Virginia, USA), using a pro- cedure based on that described by Wong et al. (1986) and the 17 lectins listed in Table I. For that, equal volumes (0.025 ml) of bacterial cells suspensions (2 X 10 8 bacteria/ml) and lectin solutions (0.24 to 1000 µg/ml) were mixed and incubated at room temperature or at 4°C for 1 and 2 hr, respectively. Cell agglutination was scored by visual inspection.Extraction and purification of the capsular polysaccharide -Capsular polysaccharide were extracted as previously described (Lloyd 1970). Briefly, strains were grown in 250 ml of BHI and, after centrifugation, 3 vol of ethanol were added to the supernatant to precipitate the polysaccharides. Precipitates were redissolved in water and reprecipitated three times with ethanol. The crude polymers were fractionated by stepwise precipitation with hexadecyltrimethylammonium bromide (Cetavlon) at pH 7.0 (Fraction A) and Cetavlonborate at pH 8.8 (Fraction B). Polysaccharides ...

Helical Disposition of Proteins and Lipopolysaccharide in the Outer Membrane ofEscherichia coli

“…Consistent with the results presented here, Stoitsova et al did not observe binding of gold-labeled ConA to E. coli K-12 (57). Thus, ConA may bind with low avidity to carbohydrate residues other than those of the O side chain, as is suggested by the lectin's ability to agglutinate E. coli serotypes having different O antigens (32). In any event, the ability to retain a substantial, visibly fluorescent ConA-linked dye was limited to cells expressing the O8 antigen, which contains repeated units of ␣-mannose (5).…”

Surface conditioning with <em>Escherichia coli</em> cell wall components can reduce biofilm formation by decreasing initial adhesion

“…Mannose is a sugar monomer which is typically found on the walls of bacterial cells [28]. Furthermore, it is one of the predominant monosaccharide components detected on the cell surfaces of various enteropathogenic E. coli serotypes [29]. The two saturated fatty acids palmitic acid (C16:0) and myristic acid (C14:0) are the dominant ones in the E. coli cell wall [30],[31], consisting of more than 50% of the fatty acid content in continuous cultures [31].…”

Virulence factors of septicemic Escherichia coli strains

“…Another virulence factor that varies between the strains is the capsule. While O2 strains possess a K-1 polysaccharide capsule, O78 strains lack such capsule, but are enveloped by a type IV capsule (KrallmannWenzel and Schmidt, 1994;Mangia et al, 1999;Ron et al, unpublished results). All the septicemic strains carry a plasmid encoding Colicin V (ColV plasmid), known to encode also the aerobactin iron uptake system as well as serum resistance (Valvano and Crosa, 1984;Crosa, 1986, 1991;Zgur-Bertok et al, 1990).…”